Mono- or di-substituted indole derivatives as dengue viral replication inhibitors

ABSTRACT

The present invention relates to mono- or di-substituted indole compounds, methods to prevent or treat dengue viral infections by using the compounds and also relates to use of the compounds as a medicine, more preferably for use as a medicine to treat or prevent dengue viral infections. The present invention furthermore relates to pharmaceutical compositions or combination preparations of the compounds, to the compositions or preparations for use as a medicine, more preferably for the prevention or treatment of dengue viral infections. The invention also relates to processes for preparation of the compounds.

RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.15/930,738, filed May 13, 2020, which is a continuation of U.S. patentapplication Ser. No. 15/571,930, filed Nov. 6, 2017, which is a 35U.S.C. § 371 nationalization of PCT application PCT/EP2016/059975 filedMay 4, 2016, which claims priority to European Patent Application15166900.9 filed May 8, 2015 and European Patent Application 16163342.5filed Mar. 31, 2016.

The present invention relates to mono- or di-substituted indolecompounds, methods to prevent or treat dengue viral infections by usingsaid compounds and also relates to said compounds for use as a medicine,more preferably for use as a medicine to treat or prevent dengue viralinfections. The present invention furthermore relates to pharmaceuticalcompositions or combination preparations of the compounds, to thecompositions or preparations for use as a medicine, more preferably forthe prevention or treatment of dengue viral infections. The inventionalso relates to processes for preparation of the compounds.

BACKGROUND OF THE INVENTION

Flaviviruses, which are transmitted by mosquitoes or ticks, causelife-threatening infections in man, such as encephalitis and hemorrhagicfever. Four distinct, but closely related serotypes of the flavivirusdengue are known, so-called DENV-1, -2, -3, and -4. Dengue is endemic inmost tropical and sub-tropical regions around the world, predominantlyin urban and semi-urban areas. According to the World HealthOrganization (WHO), 2.5 billion people of which 1 billion children areat risk of DENV infection (WHO, 2002). An estimated 50 to 100 millioncases of dengue fever [DF], half a million cases of severe denguedisease (i.e. dengue hemorrhagic fever [DHF] and dengue shock syndrome[DSS]), and more than 20,000 deaths occur worldwide each year. DHF hasbecome a leading cause of hospitalization and death amongst children inendemic regions. Altogether, dengue represents the most common cause ofarboviral disease. Because of recent large outbreaks in countriessituated in Latin America, South-East Asia and the Western Pacific(including Brazil, Puerto Rico, Venezuela, Cambodia, Indonesia, Vietnam,Thailand), numbers of dengue cases have risen dramatically over the pastyears. Not only is the number of dengue cases increasing as the diseaseis spreading to new areas, but the outbreaks tend to be more severe.

To prevent and/or control the disease associated with dengue viralinfection, the only available methods at present are mosquitoeradication strategies to control the vector. Although progress is beingmade in the development of vaccines against dengue, many difficultiesare encountered. These include the existence of a phenomenon referred toas antibody-dependent enhancement (ADE). Recovery from an infection byone serotype provides lifelong immunity against that serotype butconfers only partial and transient protection against a subsequentinfection by one of the other three serotypes. Following infection withanother serotype, pre-existing heterologous antibodies form complexeswith the newly infecting dengue virus serotype but do not neutralize thepathogen. Instead, virus entry into cells is believed to be facilitated,resulting in uncontrolled virus replication and higher peak viraltiters. In both primary and secondary infections, higher viral titersare associated with more severe dengue disease. Since maternalantibodies can easily pass on to infants by breast feeding, this mightbe one of the reasons that children are more affected by severe denguedisease than adults.

In locations with two or more serotypes circulating simultaneously, alsoreferred to as hyper endemic regions, the risk of serious dengue diseaseis significantly higher due to an increased risk of experiencing asecondary, more severe infection. Moreover, in a situation ofhyper-endemicity, the probability of the emergence of more virulentstrains is increased, which in turn augments the probability of denguehemorrhagic fever (DHF) or dengue shock syndrome.

The mosquitoes that carry dengue, including Aedes aegypti and Aedesalbopictus (tiger mosquito), are moving north on the globe. According tothe United States (US) Centers for Disease Control and Prevention (CDC),both mosquitoes are currently omnipresent in southern Texas. The spreadnorth of dengue-carrying mosquitoes is not confined to the US, but hasalso been observed in Europe.

Recently (December 2015), the dengue vaccine produced by Sanofi Pasteurwas first approved in Mexico. The vaccine has also been approved inBrazil, The Philippines and El Salvador. Regulatory review processes arecontinuing in other countries where dengue is a public health priority.Nevertheless, the vaccine leaves considerable room for improvement dueto limited efficacy, especially against DENV-1 and -2, low efficacy inflavivirus-naïve subjects and the lengthy dosing schedule.

Despite these shortcomings, the vaccine is a game changer in endemicsettings as it will offer protection to a large part of the population,but likely not to very young infants, who bear the largest burden ofdengue. In addition, the dosing schedule and very limited efficacy inflavivirus-naïve subjects make it unsuitable and likely notworthwhile/cost-effective for travelers from non-endemic areas todengue-endemic areas. The above mentioned shortcomings of the denguevaccines are the reason why there is a need for a pre-exposureprophylactic dengue antiviral.

Furthermore, today, specific antiviral drugs for the treatment orprevention of dengue fever virus infection are not available. Clearly,there is still a great unmet medical need for therapeutics for theprevention or treatment of viral infections in animals, more inparticular in humans and especially for viral infections caused byFlaviviruses, more in particular Dengue virus. Compounds with goodanti-viral potency, no or low levels of side-effects, a broad spectrumactivity against multiple Dengue virus serotypes, a low toxicity and/orgood pharmacokinetic or -dynamic properties are highly needed.

The present invention now provides compounds, mono- or di-substitutedindole derivatives, which show high potent activity against all four (4)serotypes of the Dengue virus. Also the compounds according to theinvention possess a good pharmacokinetic profile and surprisingly thesespecific compounds show an improved chiral stability.

SUMMARY OF THE INVENTION

The present invention is based on the unexpected finding that at leastone of the above-mentioned problems can be solved by the currentcompounds of the invention.

The present invention provides compounds which have been shown topossess potent antiviral activity against all four (4) serotypescurrently known. The present invention furthermore demonstrates thatthese compounds efficiently inhibit proliferation of Dengue virus(DENV). Therefore, these compounds constitute a useful class of potentcompounds that can be used in the treatment and/or prevention of viralinfections in animals, mammals and humans, more specifically for thetreatment and/or prevention of infections with Dengue viruses.

The present invention furthermore relates to the use of such compoundsas medicines and to their use for the manufacture of medicaments fortreating and/or preventing viral infections, in particular with virusesbelonging to the family of the Dengue viruses in animals or mammals,more in particular in humans. The invention also relates to methods forthe preparation of all such compounds and to pharmaceutical compositionscomprising them in an effective amount.

The present invention also relates to a method of treatment orprevention of dengue viral infections in humans by the administration ofan effective amount of one or more such compounds, or a pharmaceuticallyacceptable salt thereof optionally in combination with one or more othermedicines, like another antiviral agent or dengue vaccine or both, to apatient in need thereof.

One aspect of the invention is the provision of compounds of formula (I)

-   -   a stereo-isomeric form, a pharmaceutically acceptable salt,        solvate or polymorph thereof comprising a mono- or        di-substituted indole group; said compound is selected from the        group wherein:    -   R₁ is H, R₂ is F and R₃ is H or CH₃,    -   R₁ is H, CH₃ or F, R₂ is OCH₃ and R₃ is H,    -   R₁ is H, R₂ is OCH₃ and R₃ is CH₃,    -   R₁ is CH₃, R₂ is F and R₃ is H,    -   R₁ is CF₃ or OCF₃, R₂ is H and R₃ is H,    -   R₁ is OCF₃, R₂ is OCH₃ and R₃ is H and    -   R₁ is OCF₃, R₂ is H and R₃ is CH₃.

In particular the compounds of the invention or their stereo-isomericform, a pharmaceutically acceptable salt, solvate or polymorph thereofare selected from the group:

Another aspect of the invention is the use of a compound represented bythe following structural formula (I)

-   -   a stereo-isomeric form, a pharmaceutically acceptable salt,        solvate or polymorph thereof comprising a mono- or        di-substituted indole group; said compound is selected from the        group wherein:    -   R₁ is H, R₂ is F and R₃ is H or CH₃,    -   R₁ is H, CH₃ or F, R₂ is OCH₃ and R₃ is H and    -   R₁ is H, R₂ is OCH₃ and R₃ is CH₃,    -   R₁ is CH₃, R₂ is F and R₃ is H,    -   R₁ is CF₃ or OCF₃, R₂ is H and R₃ is H,    -   R₁ is OCF₃, R₂ is OCH₃ and R₃ is H and    -   R₁ is OCF₃, R₂ is H and R₃ is CH₃    -   for inhibiting the replication of dengue virus(es) in a        biological sample or patient.

Part of the current invention is also a pharmaceutical compositioncomprising a compound of formula (I) or a stereo-isomeric form, apharmaceutically acceptable salt, solvate or polymorph thereof togetherwith one or more pharmaceutically acceptable excipients, diluents orcarriers.

Pharmaceutically acceptable salts of the compounds of formula (I)include the acid addition and base salts thereof. Suitable acid additionsalts are formed from acids which form non-toxic salts. Suitable basesalts are formed from bases which form non-toxic salts.

The compounds of the invention may also exist in un-solvated andsolvated forms. The term “solvate” is used herein to describe amolecular complex comprising the compound of the invention and one ormore pharmaceutically acceptable solvent molecules, for example,ethanol.

The term “polymorph” refers to the ability of the compound of theinvention to exist in more than one form or crystal structure.

The compounds of the present invention may be administered ascrystalline or amorphous products. They may be obtained for example assolid plugs, powders, or films by methods such as precipitation,crystallization, freeze drying, spray drying, or evaporative drying.They may be administered alone or in combination with one or more othercompounds of the invention or in combination with one or more otherdrugs. Generally, they will be administered as a formulation inassociation with one or more pharmaceutically acceptable excipients. Theterm “excipient” is used herein to describe any ingredient other thanthe compound(s) of the invention. The choice of excipient dependslargely on factors such as the particular mode of administration, theeffect of the excipient on solubility and stability, and the nature ofthe dosage form.

The compounds of the present invention or any subgroup thereof may beformulated into various pharmaceutical forms for administrationpurposes. As appropriate compositions there may be cited allcompositions usually employed for systemically administering drugs. Toprepare the pharmaceutical compositions of this invention, an effectiveamount of the particular compound, optionally in addition salt form, asthe active ingredient is combined in intimate admixture with apharmaceutically acceptable carrier, which carrier may take a widevariety of forms depending on the form of preparation desired foradministration. These pharmaceutical compositions are desirably inunitary dosage form suitable, for example, for oral or rectaladministration. For example, in preparing the compositions in oraldosage form, any of the usual pharmaceutical media may be employed suchas, for example, water, glycols, oils, alcohols and the like in the caseof oral liquid preparations such as suspensions, syrups, elixirs,emulsions, and solutions; or solid carriers such as starches, sugars,kaolin, diluents, lubricants, binders, disintegrating agents and thelike in the case of powders, pills, capsules, and tablets. Because oftheir ease in administration, tablets and capsules represent the mostadvantageous oral dosage unit forms, in which case solid pharmaceuticalcarriers are obviously employed. Also included are solid formpreparations that can be converted, shortly before use, to liquid forms.

It is especially advantageous to formulate the aforementionedpharmaceutical compositions in unit dosage form for ease ofadministration and uniformity of dosage. Unit dosage form as used hereinrefers to physically discrete units suitable as unitary dosages, eachunit containing a predetermined quantity of active ingredient calculatedto produce the desired therapeutic effect in association with therequired pharmaceutical carrier. Examples of such unit dosage forms aretablets (including scored or coated tablets), capsules, pills, powderpackets, wafers, suppositories, injectable solutions or suspensions andthe like, and segregated multiples thereof.

Those of skill in the treatment of infectious diseases will be able todetermine the effective amount from the test results presentedhereinafter. In general it is contemplated that an effective dailyamount would be from 0.01 mg/kg to 50 mg/kg body weight, more preferablyfrom 0.1 mg/kg to 10 mg/kg body weight. It may be appropriate toadminister the required dose as two, three, four or more sub-doses atappropriate intervals throughout the day. Said sub-doses may beformulated as unit dosage forms, for example, containing 1 to 1000 mg,and in particular 5 to 200 mg of active ingredient per unit dosage form.

The exact dosage and frequency of administration depends on theparticular compound of formula (I) used, the particular condition beingtreated, the severity of the condition being treated, the age, weightand general physical condition of the particular patient as well asother medication the individual may be taking, as is well known to thoseskilled in the art. Furthermore, it is evident that the effective amountmay be lowered or increased depending on the response of the treatedsubject and/or depending on the evaluation of the physician prescribingthe compounds of the instant invention. The effective amount rangesmentioned above are therefore only guidelines and are not intended tolimit the scope or use of the invention to any extent.

The present disclosure is also intended to include any isotopes of atomspresent in the compounds of the invention. For example, isotopes ofhydrogen include tritium and deuterium and isotopes of carbon includeC-13 and C-14. The present compounds used in the current invention mayalso exist in their stereo-chemically isomeric form, defining allpossible compounds made up of the same atoms bonded by the same sequenceof bonds but having different three-dimensional structures, which arenot interchangeable. Unless otherwise mentioned or indicated, thechemical designation of compounds encompasses the mixture of allpossible stereo-chemically isomeric forms, which said compounds mightpossess.

Said mixture may contain all dia-stereomers and/or enantiomers of thebasic molecular structure of said compound. All stereo-chemicallyisomeric forms of the compounds used in the present invention either inpure form or in admixture with each other are intended to be embracedwithin the scope of the present invention including any racemic mixturesor racemates.

Pure stereoisomeric forms of the compounds and intermediates asmentioned herein are defined as isomers substantially free of otherenantiomeric or diastereomeric forms of the same basic molecularstructure of said compounds or intermediates. In particular, the term‘stereoisomerically pure’ concerns compounds or intermediates having astereoisomeric excess of at least 80% (i. e. minimum 90% of one isomerand maximum 10% of the other possible isomers) up to a stereoisomericexcess of 100% (i.e. 100% of one isomer and none of the other), more inparticular, compounds or intermediates having a stereoisomeric excess of90% up to 100%, even more in particular having a stereoisomeric excessof 94% up to 100% and most in particular having a stereoisomeric excessof 97% up to 100%. The terms ‘enantiomerically pure’ and‘diastereomerically pure’ should be understood in a similar way, butthen having regard to the enantiomeric excess, respectively thediastereomeric excess of the mixture in question.

Pure stereoisomeric forms of compounds and intermediates used in thisinvention may be obtained by the application of art-known procedures.For instance, enantiomers may be separated from each other by theselective crystallization of their diastereomeric salts with opticallyactive acids or bases. Examples thereof are tartaric acid,dibenzoyltartaric acid, ditoluoyltartaric acid and camphosulfonic acid.Alternatively, enantiomers may be separated by chromatographictechniques using chiral stationary phases. Said pure stereochemicallyisomeric forms may also be derived from the corresponding purestereochemically isomeric forms of the appropriate starting materials,provided that the reaction occurs stereospecifically. Preferably, if aspecific stereoisomer is desired, said compound will be synthesized bystereospecific methods of preparation. These methods will advantageouslyemploy enantiomerically pure starting materials.

General Synthetic Approaches

The synthesis of compounds of general formula I can be performed asoutlined in Scheme 1. 2-(4-Chloro-2-methoxyphenyl)acetic acid (II) canbe converted to the corresponding 2-(4-chloro-2-methoxyphenyl)acetylchloride (III) with a chlorination reagent like for example thionylchloride. The Friedel-Crafts reaction of the acid chloride III with asubstituted indole of general formula IV can be performed using a Lewisacid reagent like for example Et₂AlCl or TiCl₄ in a suitable solventlike for example CH₂Cl₂ or 1,2-dichloroethane, and under suitablereaction conditions that typically (but not exclusively) involvecooling, to provide the 3-acylated indole of general formula V. Theintroduction of an aniline moiety in alpha position to the carbonylmoiety of the compounds of general formula V can be accomplished by areaction sequence that involves for example bromination of V with areagent like for example phenyltrimethylammonium tribromide in asuitable solvent like for example THF, to provide the compounds ofgeneral formula VI, and subsequent reaction of the compounds of generalformula VI with 3-methoxy-5-(methyl-sulfonyl)aniline (VII) in a suitablesolvent like for example CH₃CN, and typically using a base like forexample TEA or DIPEA, to provide the compounds of general formula I asracemic mixtures. Chiral separation of the compounds of general formulaI can be performed by for example chiral chromatography to provide theEnantiomers A and B of general formula I.

In some cases, the synthesis of the intermediate of general formula Vvia the Friedel-Crafts synthesis approach, benefits from the presence ofa protecting group (PG) at the indole-N during the Friedel-Craftsreaction step, as outlined in Scheme 2. To this end, the substitutedindole of general formula IV can be converted first to an N-protectedintermediate of general formula VIII, such as for example an N-Tosylatedintermediate of general formula VIII (PG=Ts), using a reagent like forexample tosyl chloride, in the presence of a base like for examplesodium hydride. The Friedel-Crafts reaction of the substituted indole ofgeneral formula IV with acid chloride III can be performed using a Lewisacid reagent like for example Et₂AlCl or TiCl₄ in a suitable solventlike for example CH₂Cl₂ or 1,2-dichloroethane, and under suitablereaction conditions that typically (but not exclusively) involvecooling, to provide the 3-acylated N-protected indole of general formulaIX. Removal of the indole-N protecting group PG of the intermediate ofgeneral formula IX can be accomplished with a reagent like for exampleLiOH (for PG=Ts) in a solvent mixture like for example THF/water an at asuitable reaction temperature, to provide the 3-acylated indole ofgeneral formula V.

As an alternative approach, the intermediate of general formula V canalso be prepared as outlined in Scheme 3: The N-Boc-protectedsubstituted indole-3-carbaldehyde of general formula X can be convertedto the corresponding Strecker-type of intermediate of general formula XIby reaction with morpholine in the presence of reagents like for examplesodium cyanide and sodium bisulfite and in a suitable solvent like forexample a mixture of water and a water-mixable organic solvent like forexample dioxane. Alkylation of the compound of general formula XI with4-chloro-2-methoxy-benzylchloride can be accomplished in the presence ofa base like for example potassium hexamethyldisilazane and in a suitablesolvent like for example DMF to provide the compound of general formulaXII. Submission of the compound of general formula XII to a suitableaqueous acidic hydrolytic condition like for example by treatment withan aqueous hydrochloric acid solution at elevated temperature, providesthe intermediate of general formula V.

EXAMPLES

LC/MS Methods

The High Performance Liquid Chromatography (H PLC) measurement wasperformed using a LC pump, a diode-array (DAD) or a UV detector and acolumn as specified in the respective methods. If necessary, additionaldetectors were included (see table of methods below).

Flow from the column was brought to the Mass Spectrometer (MS) which wasconfigured with an atmospheric pressure ion source. It is within theknowledge of the skilled person to set the tune parameters (e.g.scanning range, dwell time . . . ) in order to obtain ions allowing theidentification of the compound's nominal monoisotopic molecular weight(MW). Data acquisition was performed with appropriate software.

Compounds are described by their experimental retention times (R_(t))and ions. If not specified differently in the table of data, thereported molecular ion corresponds to the [M+H]⁺ (protonated molecule)and/or [M−H]⁻ (deprotonated molecule). In case the compound was notdirectly ionizable the type of adduct is specified (i.e. [M+NH₄]⁺,[M+HCOO]⁻, etc. . . . ). For molecules with multiple isotopic patterns(Br, Cl), the reported value is the one obtained for the lowest isotopemass. All results were obtained with experimental uncertainties that arecommonly associated with the method used.

Hereinafter, “SQD” means Single Quadrupole Detector, “MSD” MassSelective Detector, “RT” room temperature, “BEH” bridgedethylsiloxane/silica hybrid, “DAD” Diode Array Detector, “HSS” HighStrength silica.

LCMS Method codes (Flow expressed in mL/min; column temperature (T) in °C.; Run time in minutes)

Run Method Flow time code Instrument Column Mobile phase Gradient Col T(min) LC-A Waters: Waters: A: 10 mM From 95% A to 0.8 2 Acquity ® BEHC18 CH₃COONH₄ 5% A in 1.3 min, mL/min UPLC ® - (1.7 μm, in 95% H₂O heldfor 0.7 min 55° C. DAD-SQD 2.1 × 50 mm) 5% CH₃CN B: CH₃CN LC-B Waters:Waters: A: 10 mM From 100% A to 0.7 3.5 Acquity ® HSS T3 CH₃COONH₄ 5% Ain 2.10 min, mL/min UPLC ® - (1.8 μm, in 95% H₂O + to 0% A in 0.90 min,55° C. DAD-SQD 2.1 × 100 mm) 5% CH₃CN to 5% A in 0.5 min B: CH₃CN LC-CWaters: Waters: A: 95% 84.2% A for 0.343 6.2 Acquity ® BEH C18 CH₃COONH₄0.49 min, to 10.5% mL/min UPLC ® - DAD- (1.7 μm, 7 mM/5% A in 2.18 min,40° C. Quattro 2.1 × 100 mm) CH₃CN, held for 1.94 min, Micro ™ B: CH₃CNback to 84.2% A in 0.73 min, held for 0.73 min LC-D Waters: Waters: A:10 mM From 50% A to 0.5 5 Acquity ® BEH C18 CH₃COONH₄ 10% A in 3.5 min,mL/min UPLC ® - DAD- (1.7 μm, (adjusted at held for 1.5 min 40° C.Acquity ® TQ 2.1 × 50 mm) pH 10) detector B: CH₃CN

SFC-MS Methods

The SFC measurement was performed using an Analytical Supercriticalfluid chromatography (SFC) system composed by a binary pump fordelivering carbon dioxide (CO2) and modifier, an autosampler, a columnoven, a diode array detector equipped with a high-pressure flow cellstanding up to 400 bars. If configured with a Mass Spectrometer (MS) theflow from the column was brought to the (MS). It is within the knowledgeof the skilled person to set the tune parameters (e.g. scanning range,dwell time . . . ) in order to obtain ions allowing the identificationof the compound's nominal monoisotopic molecular weight (MW). Dataacquisition was performed with appropriate software.

Analytical SFC-MS Methods (Flow expressed in mL/min; column temperature(T) in ° C.; Run time in minutes, Backpressure (BPR) in bars.

Run Method mobile Flow time code column phase gradient Col T BPR SFC-AWHELK-O1 (S, S) 5 μm A: CO₂ 50% B hold 3 7 250 × 4.6 mm Regis B: MeOH 7min, 35 100 SFC-B Daicel Chiralpak ® A: CO₂ 40% B hold 3 7 IC-H columnB: MeOH 7 min, 35 100 (5 μm, 150 × 4.6 mm) SFC-C WHELK-O1 (S, S) 5 μm A:CO₂ 60% B hold 3 9 250 × 4.6 mm Regis B: MeOH 9 min, 35 100 SFC-D DaicelChiralpak ® A: CO₂ 50% B hold 3 7 IA-H column B: MeOH 7 min, 35 100 (5μm, 250 × 4.6 mm) SFC-E Daicel Chiralpak ® A: CO₂ 10%-50% B 2.5 9.5 AS3column B: EtOH + in 6 min, 40 110 (3.0 μm, 150 × 4.6 mm) 0.2% iPrNH₂ +hold 3.5 min 3% H₂O SFC-F Daicel Chiralpak ® A: CO₂ 30% B hold 3 7 AD-Hcolumn B: iPrOH + 7 min 35 100 (5.0 μm, 150 × 4.6 mm) 0.3% iPrNH₂

Melting Points

Values are either peak values or melt ranges, and are obtained withexperimental uncertainties that are commonly associated with thisanalytical method.

DSC823e (Indicated as DSC)

For a number of compounds, melting points were determined with a DSC823e(Mettler-Toledo). Melting points were measured with a temperaturegradient of 10° C./minute. Maximum temperature was 300° C.

Optical Rotations:

Optical rotations were measured on a Perkin-Elmer 341 polarimeter with asodium lamp and reported as follows: [α]° (λ, c g/100 ml, solvent, T°C.).

[α]_(λ) ^(T)=(100α)/(l×c): where l is the path length in dm and c is theconcentration in g/100 ml for a sample at a temperature T (° C.) and awavelength λ (in nm). If the wavelength of light used is 589 nm (thesodium D line), then the symbol D might be used instead. The sign of therotation (+ or −) should always be given. When using this equation theconcentration and solvent are always provided in parentheses after therotation. The rotation is reported using degrees and no units ofconcentration are given (it is assumed to be g/100 ml).

Example 1: Synthesis of2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)ethanone(Compound 1) and chiral separation into Enantiomers 1A and 1B

Synthesis of Intermediate 1a

2-(4-Chloro-2-methoxyphenyl)acetic acid [CAS 170737-95-8] (5.8 g, 28.9mmol) was added in small portions to thionyl chloride (50 mL) and theresulting solution was stirred overnight at 60° C. The solvent wasconcentrated under reduced pressure and co-evaporated with toluene togive 2-(4-chloro-2-methoxyphenyl)-acetyl chloride 1a (6.5 g) as an oilyresidue that was used without further purification in the next step.

Synthesis of Intermediate 1b

Diethylaluminum chloride 1M in hexane (37.1 mL, 37.1 mmol) was addeddropwise at 0° C. to a solution of 6-fluoro-1H-indole [CAS 399-51-9](3.34 g, 24.76 mmol) in CH₂Cl₂ (100 mL). After 30 min at 0° C., asolution of 2-(4-chloro-2-methoxyphenyl)acetyl chloride 1a (6.3 g, 28.76mmol) in CH₂Cl₂ (100 mL) was added slowly at 0° C. The reaction wasstirred at 0° C. for 3 h. Ice-water was added and the precipitate wasfiltered off, washed with water and a small amount of CH₂Cl₂. The solidswere dried under vacuum at 70° C. overnight to give2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-1H-indol-3-yl)ethanone 1b (4.9g).

Synthesis of Intermediate 1c

At 0° C., a solution of phenyltrimethylammonium tribromide [CAS4207-56-1] (5.8 g, 15.4 mmol) in THF (65 mL) was added dropwise to amixture of2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-1H-indol-3-yl)ethanone 1b (4.9g, 15.4 mmol) in THF (60 mL). The mixture was stirred at 0° C. for 1 hand at room temperature for 2.5 h. The precipitate was filtered off andwashed with EtOAc. The combined filtrates were concentrated underreduced pressure. The residue was taken up with EtOAc and washed withwater. A precipitate appeared in the organic layer and was filtered offand dried to provide a first batch of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-1H-indol-3-yl)ethanone1c (4.6 g). The organic layer was separated, dried over MgSO₄, filteredand the solvent was evaporated under reduced pressure. The residue wascrystallized from EtOAc, the precipitate was filtered off, washed withEt₂O and dried under vacuum to provide a second fraction of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-1H-indol-3-yl)ethanone1c (1.6 g).

Synthesis of Compound 1 and Chiral Separation of Enantiomers 1A and 1B

A mixture of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-1H-indol-3-yl)-ethanone1c (3 g, 7.56 mmol), 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (2.28 g, 11.35 mmol) and diisopropylethylamine (1.95 mL,11.35 mmol) in CH₃CN (60 mL) and THF (30 mL) was stirred at 70° C. for24 h. The reaction was diluted with EtOAc. The organic layer was washedwith 1N HCl (twice) and water, dried over MgSO₄, filtered and thesolvent was concentrated under reduced pressure. The residue waspurified by flash chromatography on silica gel (15-40 μm, 80 g, Mobilephase: CH₂Cl₂/MeOH 99.5/0.5). A second purification was carried out byflash chromatography on silica gel (15-40 μm, 80 g, Mobile phase:CH₂Cl₂/MeOH 99.7/0.3). The pure fractions were combined and concentratedunder reduced pressure to give2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)ethanone(Compound 1, 2 g) as a racemic mixture.

The enantiomers of Compound 1 were separated via Chiral SFC (Stationaryphase: Chiralpak® AD-H 5 μm 20×250 mm, Mobile phase: 50% CO₂, 50% MeOH)yielding 740 mg of the first eluted enantiomer and 720 mg of the secondeluted enantiomer. The first eluted enantiomer was crystallized fromCH₃CN/Et₂O. The precipitate was filtered off and dried to giveEnantiomer 1A (645 mg). The second eluted enantiomer was crystallizedfrom CH₃CN/Et₂O. The precipitate was filtered off and dried to giveEnantiomer 1B (632 mg).

Compound 1:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 4.00 (s, 3H)6.24 (d, J=7.9 Hz, 1H) 6.58 (s, 2H) 6.91 (s, 1H) 6.97 (dd, J=8.7, 1.9Hz, 1H) 7.02-7.09 (m, 2H) 7.12 (d, J=1.9 Hz, 1H) 7.27 (dd, J=9.5, 1.9Hz, 1H) 7.35 (d, J=8.5 Hz, 1H) 8.14 (dd, J=8.7, 5.5 Hz, 1H) 8.44 (s, 1H)12.10 (br. s., 1H)

LC/MS (method LC-C): R_(t) 3.08 min, MH⁺517

Melting point: 174° C.

Enantiomer 1A:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 4.00 (s, 3H)6.24 (d, J=7.9 Hz, 1H) 6.59 (s, 2H) 6.91 (s, 1H) 6.97 (dd, J=8.8, 2.2Hz, 1H) 7.02-7.10 (m, 2H) 7.12 (d, J=2.2 Hz, 1H) 7.27 (dd, J=9.6, 2.2Hz, 1H) 7.35 (d, J=8.2 Hz, 1H) 8.14 (dd, J=8.8, 5.7 Hz, 1H) 8.44 (s, 1H)12.10 (br. s., 1H)

LC/MS (method LC-C): R_(t) 3.09 min, MH⁺517

[α]_(D) ²⁰: +130.3° (c 0.277, DMF)

Chiral SFC (method SFC-D): R_(t) 3.41 min, MH⁺517, chiral purity 100%.

Melting point: 220° C.

Enantiomer 1B:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 4.00 (s, 3H)6.24 (d, J=7.6 Hz, 1H) 6.53-6.65 (m, 2H) 6.91 (s, 1H) 6.97 (dd, J=8.6,2.0 Hz, 1H) 7.01-7.09 (m, 2H) 7.12 (d, J=2.0 Hz, 1H) 7.27 (dd, J=9.6,2.0 Hz, 1H) 7.35 (d, J=8.1 Hz, 1H) 8.14 (dd, J=8.6, 5.6 Hz, 1H) 8.43 (s,1H) 12.09 (br. s., 1H)

LC/MS (method LC-C): R_(t) 3.09 min, MH⁺517

[α]_(D) ²⁰: −135.3° (c 0.283, DMF)

Chiral SFC (method SFC-D): R_(t) 4.89 min, MH⁺517, chiral purity 99.35%.

Melting point: 218° C.

Example 1.1: Chiral Stability of Enantiomer 1A at pH 7.4

The chiral stability of Enantiomer 1A (R=OMe) was evaluated bydetermination of the enantiomeric excess (ee %) after incubation for 24h and 48 h in a buffered solution at pH 7.4 at 40° C. and 60° C. Toassess the influence of the methoxy-substituent of Enantiomer 1A (R=OMe)on the stability against racemization, the chiral stability ofEnantiomer 1′A (R═H) was tested under the same conditions. To this end,5 μM buffered (pH=7.4) solutions of 1A and 1′A were prepared by mixing25 μL of a 100 μM solution of 1A or 1′A in DMSO with 475 μL aqueousbuffer pH 7.4. Samples were taken 24 h and 48 h after incubation at 40°C. and 60° C. The analytical samples were analyzed by Chiral SFC (MSdetection) and the chiral purity was expressed as the enantiomericexcess (ee %=% enantiomer A−% enantiomer B). Both Enantiomers 1A and 1′Ahad a chiral purity of 100% prior to their incubation.

ee % Sampling timepoints (h) Compound Temperature 24 48 1A 40° C. 100100 60° C. 95 88 1′A 40° C. 21 10 60° C. 0 0

Example 2: synthesis of2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-7-methyl-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)ethanone(Compound 2) and Chiral Separation into Enantiomers 2A and 2B

Synthesis of Intermediate 2a

Diethylaluminum chloride 1M in hexane (20 mL, 20.0 mmol) was addeddropwise at 0° C. to a solution of 6-fluoro-7-methyl-1H-indole [CAS57817-10-4] (1.50 g, 10.1 mmol) in CH₂Cl₂ (45 mL). After 30 min at 0°C., a solution of 2-(4-chloro-2-methoxyphenyl)acetyl chloride (3.30 g,15.1 mmol, synthesis: see Example 1) in dichloromethane (30 mL) wasadded slowly. The reaction mixture was stirred at 0° C. for 3 h. 1MRochelle salt solution (50 mL) was added and the reaction mixture wasstirred at room temperature for 1 h. The solids were filtered off andpartitioned between EtOAc and 1N HCl. The phases were separated. Theaqueous phase was extracted with EtOAc. The organic phases werecombined, washed with brine, dried over MgSO₄, filtered and concentratedunder reduced pressure. The residue was triturated with EtOAc andheptane. The precipitate was filtered off to give2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-7-methyl-1H-indol-3-yl)ethanone2a (2.00 g).

Synthesis of Intermediate 2b

A solution of phenyltrimethylammonium tribromide [CAS 4207-56-1] (2.49g, 6.6 mmol) in THF (45 mL) was added dropwise at 0° C. to a solution of2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-7-methyl-1H-indol-3-yl)ethanone2a (2.00 g, 6.0 mmol) in THF (65 mL). The mixture was stirred at roomtemperature overnight. The precipitate was filtered off and washed withEtOAc. The combined filtrates were concentrated under reduced pressure.The residue was taken up with a minimum of acetonitrile. The precipitatewas filtered off, washed with acetonitrile and dried under vacuum togive a first batch of2-bromo-2-(4-chloro-2-methoxy-phenyl)-1-(6-fluoro-7-methyl-1H-indol-3-yl)ethanone2b (1.51 g). The filtrate was concentrated under reduced pressure. Theresidue was taken up with a minimum of acetonitrile. The precipitate wasfiltered off, washed with acetonitrile and dried under vacuum to give asecond fraction of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-7-methyl-1H-indol-3-yl)ethanone2b (0.70 g).

Synthesis of Compound 2 and Chiral Separation of Enantiomers 2A and 2B

A mixture of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-7-methyl-1H-indol-3-yl)ethanone2b (1.8 g, 4.36 mmol) and 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (2.6 g, 13.0 mmol) in THF (9 mL) and CH₃CN (9 mL) was heatedat 100° C. under microwave irradiation for 50 min. The reaction mixturewas diluted with EtOAc and washed with 1N HCl. The phases wereseparated. The organic phase was washed with an aqueous saturated NaHCO₃solution and brine, dried over MgSO₄, filtered and concentrated underreduced pressure. The residue was taken up with a minimum ofacetonitrile. The precipitate was filtered off, washed with acetonitrileand dried under vacuum to give2-(4-chloro-2-methoxy-phenyl)-1-(6-fluoro-7-methyl-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)-phenyl)amino)ethanone(Compound 2, 1.7 g) as a racemic mixture.

The chiral separation of the enantiomers of Compound 2 (1.59 g) wasperformed via Preparative SFC (Stationary phase: (S,S)-Whelk-O1 5 μm250×21.1 mm, Mobile phase: 50% CO₂, 50% MeOH). The product fractionswere combined and evaporated under reduced pressure. The first elutedenantiomer (746 mg) was further purified by column chromatography onsilica gel (15-40 μm, 24 g, Mobile phase: CH₂Cl₂/MeOH 99.5/0.5). Thepure fractions were combined and evaporated under reduced pressure (560mg). The residue was solidified by trituration with a mixture of Et₂Oand a few drops of CH₃CN. The solids were filtered off and dried undervacuum to give Enantiomer 2A (473 mg). The second eluted enantiomer (732mg) was further purified by column chromatography over silica gel (15-40μm, 24 g, Mobile phase: CH₂Cl₂/MeOH 99.5/0.5). The pure fractions werecombined and evaporated under reduced pressure (550 mg). The residue wassolidified by trituration with a mixture of Et₂O and a few drops ofCH₃CN. The solids were filtered off and dried under vacuum to give ofEnantiomer 2B (457 mg).

Compound 2:

¹H NMR (300 MHz, DMSO-d₆) δ ppm 2.38 (d, J=1.5 Hz, 3H) 3.10 (s, 3H) 3.73(s, 3H) 4.01 (s, 3H) 6.27 (d, J=7.9 Hz, 1H) 6.55-6.63 (m, 2H) 6.93 (m,1H) 6.94-7.09 (m, 3H) 7.13 (d, J=1.9 Hz, 1H) 7.35 (d, J=8.3 Hz, 1H) 7.97(dd, J=8.7, 5.3 Hz, 1H) 8.45 (s, 1H) 12.23 (br. s, 1H)

LC/MS (method LC-D): R_(t) 1.68 min, MH⁺531

Enantiomer 2A:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 2.37-2.39 (m, 3H) 3.09 (s, 3H) 3.72 (s,3H) 4.01 (s, 3H) 6.26 (d, J=7.9 Hz, 1H) 6.54-6.63 (m, 2H) 6.92 (s, 1H)6.97 (dd, J=8.4, 1.9 Hz, 1H) 7.02 (dd, J=9.9, 9.0 Hz, 1H) 7.07 (d, J=7.9Hz, 1H) 7.13 (d, J=1.9 Hz, 1H) 7.35 (d, J=8.4 Hz, 1H) 7.96 (dd, J=8.5,5.4 Hz, 1H) 8.45 (s, 1H) 12.24 (br. s., 1H)

LC/MS (method LC-C): R_(t) 3.20 min, MH⁺531

[α]_(D) ²⁰: +104.5° (c 0.2545, DMF)

Chiral SFC (method SFC-A): R_(t) 4.22 min, MH⁺531, chiral purity 100%.

Enantiomer 2B:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 2.36-2.41 (m, 3H) 3.09 (s, 3H) 3.72 (s,3H) 4.01 (s, 3H) 6.26 (d, J=7.9 Hz, 1H) 6.57-6.64 (m, 2H) 6.92 (s, 1H)6.97 (dd, J=8.2, 1.9 Hz, 1H) 6.99-7.04 (m, 1H) 7.07 (d, J=7.9 Hz, 1H)7.13 (d, J=1.9 Hz, 1H) 7.35 (d, J=8.2 Hz, 1H) 7.96 (dd, J=8.7, 5.2 Hz,1H) 8.45 (s, 1H) 12.24 (br. s., 1H)

LC/MS (method LC-C): R_(t) 3.20 min, MH⁺531

[α]_(D) ²⁰: −104.1° (c 0.2536, DMF)

Chiral SFC (method SFC-A): R_(t) 5.12 min, MH⁺531, chiral purity 99.53%.

Example 3: synthesis2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)ethanone(Compound 3) and Chiral Separation into Enantiomers 3A and 3B

Synthesis of Intermediate 3a

A solution of NaHSO₃ (5.7 g, 54.5 mmol) in water (45 mL) was added to astirring solution of tert-butyl3-formyl-6-methoxy-1H-indole-1-carboxylate [CAS 847448-73-1] (10 g, 36.3mmol) in dioxane (45 mL). After 15 min, morpholine (4.8 mL, 54.5 mmol)was added and 35 min later, sodium cyanide (NaCN) (1.96 g, 40 mmol) wasadded. The resulting suspension was stirred at room temperature for 3days, until completion of the reaction. The product was filtered off andwashed with a 1/1 mixture of dioxane/water (3×35 mL), and subsequentlywith water (3×45 mL) and dried under vacuum at 60° C. The solids werestirred up in Et₂O (125 mL), filtered off, washed with Et₂O (3×) anddried under vacuum at 50° C. to provide tert-butyl3-(cyano(morpholino)methyl)-6-methoxy-1H-indole-1-carboxylate 3a (12.3g).

Synthesis of Intermediate 3b

A mixture of tert-butyl3-(cyano(morpholino)methyl)-6-methoxy-1H-indole-1-carboxylate 3a (6.0 g,16.2 mmol) in dry DMF (80 mL) was stirred under N₂-atmosphere whilecooling on an ice-bath. A solution of KHMDS 0.5 M in toluene (35.5 mL,17.8 mmol) was added dropwise over 10 min. After stirring for anadditional 10 min, 4-chloro-1-(chloromethyl)-2-methoxybenzene [CAS101079-84-9] (3.09 g, 16.2 mmol) was added and the resulting mixture wasstirred at room temperature for 20 h. The reaction mixture was pouredout into cold water (400 mL) and the product was extracted with Et₂O(2×). The combined organic layers were washed with brine, dried overMgSO₄, filtered, evaporated under reduced pressure and co-evaporatedwith xylene. The residue was purified by flash chromatography(Stationary phase: Grace Reveleris® silica 120 g, Mobile phase:heptane/EtOAc gradient 100/0 to 20/80). The desired fractions werecombined, evaporated under reduced pressure and co-evaporated withdioxane to give tert-butyl3-(2-(4-chloro-2-methoxyphenyl)-1-cyano-1-morpholinoethyl)-6-methoxy-1H-indole-1-carboxylate3b (7.75 g).

Synthesis of Intermediate 3c

To a stirred suspension of tert-butyl3-(2-(4-chloro-2-methoxyphenyl)-1-cyano-1-morpholinoethyl)-6-methoxy-1H-indole-1-carboxylate3b (7.75 g, 14.7 mmol) in dioxane (40 mL) and water (20 mL) was added asolution of HCl 6 M in isopropanol (36.8 mL, 220 mmol). The resultingmixture was stirred at 60° C. for 4 h and subsequently at 80° C. for 1h. After cooling to room temperature, the mixture was left standing for20 h to allow crystallization of the reaction product. The product wasfiltered off, washed with a 1/1/1 mixture of iPrOH/H₂O/dioxane (2×15 mL)and dried under vacuum at 50° C. to give2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-1H-indol-3-yl)ethanone 3c(3.67 g).

Synthesis of Compound 3 and Chiral Separation of Enantiomers 3A and 3B

A stirred mixture of2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-1H-indol-3-yl)-ethanone 3c (2g, 6.07 mmol) in THF (80 mL) was cooled on an ice-bath under N₂-atm.Phenyltrimethylammonium tribromide [CAS 4207-56-1] (2.39 g, 6.37 mmol)was added and the reaction mixture was stirred at 0° C. for 1 h andsubsequently at room temperature for 1.5 h.3-Methoxy-5-(methylsulfonyl)aniline [CAS 62606-02-4] (3.66 g, 18.2 mmol)was added and the solvent was evaporated under reduced pressure. Theresidue was dissolved in CH₃CN (100 mL). Diisopropylethylamine (2.09 mL,12.1 mmol) was added and the reaction mixture was heated at 55° C. for27 h. The reaction mixture was allowed to cool to room temperature andpoured out into stirring water (400 mL). The product was extracted with2-MeTHF (2×). The combined organic layers were washed with brine, driedover MgSO₄, filtered and evaporated under reduced pressure. The residue(8 g) was purified by flash chromatography (stationary phase: GraceReveleris® silica 120 g, Mobile phase: heptane/EtOAc gradient from 100/0to 0/100). The desired fractions were combined and evaporated underreduced pressure. The residue (5.4 g) was further purified byPreparative HPLC (Stationary phase: RP XBridge® Prep C18 OBD—10 μm,50×150 mm, Mobile phase: 0.25% NH₄HCO₃ solution in water, CH₃CN). Theproduct fractions were combined and evaporated under reduced pressureand subsequently co-evaporated with MeOH. The residue was crystallizedfrom a mixture of EtOAc (15 mL), CH₃CN (2 mL) and MeOH (2 mL). Thesolids were filtered off, washed with EtOAc (3×) and dried under vacuumat 50° C. to provide2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)ethanone(Compound 3, 681 mg) as a racemic mixture.

The chiral separation of the enantiomers of Compound 3 (0.63 g) wasperformed via Normal Phase Chiral separation (Stationary phase: AS 20μM, Mobile phase: 100% methanol). The product fractions were combinedand evaporated under reduced pressure. The first eluted enantiomer waspurified by flash chromatography (Stationary phase: Grace Reveleris®silica 12 g, Mobile phase: heptane/EtOAc/EtOH gradient from 100/0/0 to40/45/15). The desired fractions were combined and evaporated, andco-evaporated with EtOAc. The remaining oil was solidified by stirringup in H₂O (4 mL) and slow addition of MeOH (1.6 mL). After stirring for20 minutes, the product was filtered off, washed (3×) with a 1/2 mixtureof MeOH/H₂O and dried under vacuum at 50° C. to provide Enantiomer 3A(168 mg) as an amorphous solid. The second eluted enantiomer waspurified by flash chromatography (Stationary phase: Grace Reveleris®silica 12 g, Mobile phase: heptane/EtOAc/EtOH gradient from 100/0/0 to40/45/15). The desired fractions were combined, evaporated under reducedpressure and co-evaporated with EtOAc. The remaining foam was solidifiedby stirring up in H₂O (4 mL) and slow addition of MeOH (2 mL). Afterstirring for 15 minutes, the product was filtered off, washed (3×) witha 1/2 mixture of MeOH/H₂O and dried at 50° C. under vacuum to provideEnantiomer 3B (146 mg) as an amorphous solid.

Compound 3:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.77 (s, 3H)4.01 (s, 3H) 6.21 (d, J=7.9 Hz, 1H) 6.54-6.64 (m, 2H) 6.83 (dd, J=8.7,2.3 Hz, 1H) 6.91 (t, J=1.4 Hz, 1H) 6.94-6.99 (m, 2H) 7.04 (d, J=7.7 Hz,1H) 7.12 (d, J=2.0 Hz, 1H) 7.35 (d, J=8.1 Hz, 1H) 8.02 (d, J=8.8 Hz, 1H)8.30 (s, 1H) 11.84 (s, 1H) LC/MS (method LC-A): R_(t) 1.20 min, MH⁺529

Enantiomer 3A:

¹H NMR (360 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.77 (s, 3H)4.01 (s, 3H) 6.22 (d, J=8.1 Hz, 1H) 6.55-6.61 (m, 2H) 6.84 (dd, J=8.8,2.2 Hz, 1H) 6.91 (t, J=1.8 Hz, 1H) 6.94-7.00 (m, 2H) 7.07 (d, J=7.0 Hz,1H) 7.13 (d, J=1.8 Hz, 1H) 7.35 (d, J=8.4 Hz, 1H) 8.02 (d, J=8.8 Hz, 1H)8.32 (d, J=2.9 Hz, 1H) 11.87 (d, J=2.6 Hz, 1H)

LC/MS (method LC-A): R_(t) 1.08 min, MH⁺529

[α]_(D) ²⁰: +134.9° (c 0.545, DMF)

Chiral SFC (method SFC-E): R_(t) 4.31 min, MH⁺529, chiral purity 100%.

Enantiomer 3B:

¹H NMR (360 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.77 (s, 3H)4.01 (s, 3H) 6.21 (d, J=8.1 Hz, 1H) 6.54-6.62 (m, 2H) 6.83 (dd, J=8.6,2.4 Hz, 1H) 6.91 (t, J=1.5 Hz, 1H) 6.94-6.99 (m, 2H) 7.07 (d, J=7.0 Hz,1H) 7.13 (d, J=1.8 Hz, 1H) 7.35 (d, J=8.1 Hz, 1H) 8.02 (d, J=8.8 Hz, 1H)8.32 (d, J=2.9 Hz, 1H) 11.87 (br d, J=2.2 Hz, 1H)

LC/MS (method LC-A): R_(t) 1.08 min, MH⁺529

[α]_(D) ²⁰: −116.7° (c 0.51, DMF)

Chiral SFC (method SFC-E): R_(t) 4.63 min, MH⁺529, chiral purity 94.7%.

Example 4: Synthesis of2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methyl-sulfonyl)phenyl)amino)-1-(6-methoxy-5-methyl-1H-indol-3-yl)ethanone(Compound 4) and chiral separation into Enantiomers 4A and 4B

Synthesis of Intermediate 4a

Diethylaluminum chloride 1M in hexane (13.5 mL, 13.5 mmol) was addeddropwise at 0° C. to a solution of 6-methoxy-5-methyl-1H-indole [CAS1071973-95-9] (1.45 g, 9 mmol) in CH₂Cl₂ (45 mL). After 30 min at 0° C.,a solution of 2-(4-chloro-2-methoxyphenyl)acetyl chloride 1a (2.4 g,10.9 mmol) in CH₂Cl₂ (45 mL) was added slowly at 0° C. The reaction wasstirred at 0° C. for 3 h. Ice-water was added and the precipitate wasfiltered off and washed with water. The solid was dried under vacuum togive2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-5-methyl-1H-indol-3-yl)ethanone4a (2.1 g).

Synthesis of Intermediate 4b

At 0° C., a solution of phenyltrimethylammonium tribromide [CAS4207-56-1] (2.4 g, 6.4 mmol) in THF (65 mL) was added dropwise to amixture of2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-5-methyl-1H-indol-3-yl)ethanone4a (2.1 g, 6.1 mmol) in THF (60 mL). The mixture was stirred at 0° C.for 1 h and at room temperature for 2.5 h. The precipitate was filteredoff and washed with EtOAc. The filtrate was concentrated under reducedpressure. The residue was taken up with the minimum of diisopropylether.The precipitate was filtered off and dried under vacuum to give2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-5-methyl-1H-indol-3-yl)ethanone4b (2.36 g).

Synthesis of Compound 4 and Chiral Separation of Enantiomers 4A and 4B

A mixture of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-5-methyl-1H-indol-3-yl)ethanone4b (4.0 g, 9.46 mmol), 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (2.86 g, 14.2 mmol) and diisopropylethylamine (2.44 mL, 14.2mmol) in CH₃CN/THF (1/1) (100 mL) was stirred at 45° C. for 72 h. Thesolvents were removed under reduced pressure. The residue was dissolvedin EtOAc. The organic layer was washed twice with 1N HCl, washed withwater, dried over MgSO₄, filtered and concentrated under reducedpressure. The compound was crystallized from CH₃CN/diisopropylether togive2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)-1-(6-methoxy-5-methyl-1H-indol-3-yl)ethanone(Compound 4, 1.1 g) as a racemic mixture. The chiral separation of theenantiomers of Compound 4 was performed via Preparative Chiral SFC(Stationary phase: (S,S)-Whelk-O1 5 μm 250×21.1 mm, Mobile phase: 45%CO₂, 55% MeOH) yielding 500 mg of the first eluted enantiomer and 531 mgof the second eluted enantiomer. The first eluted enantiomer wascrystallized from CH₃CN/Et₂O to afford Enantiomer 4A (401 mg). Thesecond eluted was crystallized from CH₃CN/Et₂O to afford Enantiomer 4B(396 mg).

Compound 4:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 2.21 (s, 3H) 3.09 (s, 3H) 3.72 (s, 3H)3.79 (s, 3H) 4.01 (s, 3H) 6.20 (d, J=7.9 Hz, 1H) 6.58 (s, 2H) 6.88-6.93(m, 2H) 6.96 (dd, J=8.5, 1.9 Hz, 1H) 7.02 (d, J=7.9 Hz, 1H) 7.12 (d,J=1.9 Hz, 1H) 7.34 (d, J=8.5 Hz, 1H) 7.89 (s, 1H) 8.24 (s, 1H) 11.78(br. s., 1H)

LC/MS (method LC-C): R_(t) 3.16 min, MH⁺543

Melting point: 208° C.

Enantiomer 4A:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 2.21 (s, 3H) 3.09 (s, 3H) 3.72 (s, 3H)3.79 (s, 3H) 4.01 (s, 3H) 6.20 (d, J=7.6 Hz, 1H) 6.58 (d, J=1.6 Hz, 2H)6.87-6.93 (m, 2H) 6.96 (dd, J=8.2, 1.9 Hz, 1H) 7.02 (d, J=7.6 Hz, 1H)7.12 (d, J=1.9 Hz, 1H) 7.34 (d, J=8.2 Hz, 1H) 7.89 (s, 1H) 8.25 (s, 1H)11.78 (br. s., 1H) LC/MS (method LC-C): R_(t) 3.15 min, MH⁺543

[α]_(D) ²⁰: +141.8° (c 0.3936, DMF)

Chiral SFC (method SFC-C): R_(t) 4.95 min, MH⁺543, chiral purity 100%.

Melting point: 173° C.

Enantiomer 4B:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 2.21 (s, 3H) 3.09 (s, 3H) 3.72 (s, 3H)3.79 (s, 3H) 4.01 (s, 3H) 6.20 (d, J=7.9 Hz, 1H) 6.58 (s, 2H) 6.88-6.93(m, 2H) 6.96 (dd, J=8.2, 1.9 Hz, 1H) 7.02 (d, J=7.9 Hz, 1H) 7.12 (d,J=1.9 Hz, 1H) 7.34 (d, J=8.2 Hz, 1H) 7.90 (s, 1H) 8.25 (s, 1H) 11.79(br. s., 1H)

LC/MS (method LC-C): R_(t) 3.15 min, MH⁺543

[α]_(D) ²⁰: −142.2° (c 0.3909, DMF)

Chiral SFC (method SFC-C): R_(t) 6.84 min, MH⁺543, chiral purity 100%.

Melting point: 174° C.

Example 5: Synthesis of2-(4-chloro-2-methoxyphenyl)-1-(5-fluoro-6-methoxy-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)ethanone(Compound 5) and Chiral Separation into Enantiomers 5A and 5B

Synthesis of Intermediate 5a

Diethylaluminum chloride 1M in hexane (15.7 mL, 15.7 mmol) was addeddropwise at 0° C. to a solution of 5-fluoro-6-methoxy-1H-indole [CAS1211595-72-0] (2 g, 12.1 mmol) in CH₂Cl₂ (50 mL). After 30 min at 0° C.,a solution of 2-(4-chloro-2-methoxyphenyl)acetyl chloride 1a (3.2 g,14.6 mmol) in CH₂Cl₂ (50 mL) was added slowly at 0° C. The reaction wasstirred at 0° C. for 3 h. Ice-water was added and the precipitate wasfiltered off, washed with water and the minimum of CH₂Cl₂. The solid wasdried under vacuum to give2-(4-chloro-2-methoxyphenyl)-1-(5-fluoro-6-methoxy-1H-indol-3-yl)ethanone5a (2.82 g).

Synthesis of Intermediate 5b

At 0° C., a solution of phenyltrimethylammonium tribromide [CAS4207-56-1] (3.5 g, 8.1 mmol) in THF (20 mL) was added dropwise to asolution of2-(4-chloro-2-methoxyphenyl)-1-(5-fluoro-6-methoxy-1H-indol-3-yl)ethanone5a (2.82 g, 8.1 mmol) in THF (46 mL). The mixture was stirred at 0° C.for 1 h and at room temperature for 4 h. The precipitate was filteredoff and washed with EtOAc. The filtrate was concentrated under reducedpressure. The residue was dissolved in EtOAc and washed with water. Theorganic phase was dried over MgSO₄, filtered and the solvent wasevaporated under reduced pressure. The residue was taken up with theminimum of EtOAc. The precipitate was filtered off and dried undervacuum to give2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(5-fluoro-6-methoxy-1H-indol-3-yl)ethanone5b (2.5 g).

Synthesis of Compound 5 and Chiral Separation of Enantiomers 5A and 5B

A mixture of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(5-fluoro-6-methoxy-1H-indol-3-yl)ethanone5b (2.5 g, 5.86 mmol), 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (1.415 g, 7.03 mmol) and diisopropylethylamine (1.515 mL,8.79 mmol) in CH₃CN (55 mL) and THF (100 mL) was stirred at 50° C. for10 days. The solvents were removed under reduced pressure. The residuewas purified by flash chromatography on silica gel (15-40 μm, 80 g,Mobile phase: CH₂Cl₂/CH₃OH 99.25/0.75). The pure fractions were combinedand evaporated. The compound was dissolved in EtOAc and stirred with HCl1N for 15 min. A precipitate appeared, and was filtered off and driedunder vacuum to give2-(4-chloro-2-methoxyphenyl)-1-(5-fluoro-6-methoxy-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)-amino)ethanone(Compound 5, 1.3 g) as a racemic mixture.

The chiral separation of the enantiomers of Compound 5 was performed viaPreparative Chiral SFC (Stationary phase: Chiralpak® IC 5 μm 250×20 mm,Mobile phase: 55% CO₂, 45% MeOH). The product fractions were combinedand evaporated. The first eluted enantiomer was solidified bytrituration with heptane/diisopropylether. The solids were filtered offand dried under vacuum to provide Enantiomer 5A (502 mg) as an amorphouswhite powder. The second eluted enantiomer was solidified by triturationwith heptane/diisopropylether. The solids were filtered off and driedunder vacuum to provide Enantiomer 5B (490 mg) as an amorphous whitepowder.

Compound 5:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.85 (s, 3H)4.00 (s, 3H) 6.21 (d, J=7.9 Hz, 1H) 6.58 (d, J=1.3 Hz, 2H) 6.90 (s, 1H)6.97 (dd, J=8.2, 1.9 Hz, 1H) 7.06 (d, J=7.9 Hz, 1H) 7.10-7.18 (m, 2H)7.34 (d, J=8.2 Hz, 1H) 7.82 (d, J=12.0 Hz, 1H) 8.35 (s, 1H) 11.98 (br.s., 1H)

LC/MS (method LC-C): R_(t) 3.01 min, MH⁺547

Melting point: 182° C.

Enantiomer 5A:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.85 (s, 3H)4.00 (s, 3H) 6.21 (d, J=7.9 Hz, 1H) 6.58 (d, J=1.3 Hz, 2H) 6.90 (s, 1H)6.97 (dd, J=8.2, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.11-7.17 (m, 2H)7.34 (d, J=8.2 Hz, 1H) 7.82 (d, J=11.7 Hz, 1H) 8.35 (s, 1H) 11.98 (br.s., 1H) LC/MS (method LC-C): R_(t) 3.00 min, MH⁺547

[α]_(D) ²⁰: +136.4° (c 0.28, DMF)

Chiral SFC (method SFC-B): R_(t) 3.43 min, MH⁺547, chiral purity 100%.

Enantiomer 5B:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.85 (s, 3H)4.00 (s, 3H) 6.21 (d, J=7.9 Hz, 1H) 6.58 (d, J=1.3 Hz, 2H) 6.90 (s, 1H)6.97 (dd, J=8.2, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.11-7.19 (m, 2H)7.34 (d, J=8.2 Hz, 1H) 7.82 (d, J=11.7 Hz, 1H) 8.35 (s, 1H) 11.95 (br.s., 1H) LC/MS (method LC-C): R_(t) 3.00 min, MH⁺547 [α]_(D) ²⁰: −126.3°(c 0.2755, DMF) Chiral SFC (method SFC-B): R_(t) 4.80 min, MH⁺547,chiral purity 98.06%.

Example 6: Synthesis of2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methyl-sulfonyl)phenyl)amino)-1-(6-methoxy-7-methyl-1H-indol-3-yl)ethanone(Compound 6) and chiral separation into Enantiomers 6A and 6B

Synthesis of Intermediate 6a

Diethylaluminum chloride 1M in hexane (32.8 mL, 32.8 mmol) was addeddropwise to a cooled (−30° C.) solution of 6-methoxy-7-methyl-1H-indole[CAS 19500-05-1] (3.53 g, 21.9 mmol) in CH₂Cl₂ (150 mL). After stirringfor 15 min at −30° C., a solution of 2-(4-chloro-2-methoxyphenyl)acetylchloride 1a (6.71 g, 30.6 mmol) in CH₂Cl₂ (150 mL) was added slowly at−30° C. The reaction was stirred at −30° C. for 1 h and was allowed towarm to room temperature while stirring for 2 h. The reaction mixturewas poured out in ice-water/Rochelle salt. The mixture was filtered overa short pad of Dicalite® and the filter cake was rinsed several timeswith THF. The layers were separated. The aqueous layer was extractedwith THF. The combined organic layers were washed with brine, water,dried over MgSO₄, filtered, and evaporated under reduced pressure. Thesolid residue was suspended in CH₂Cl₂ (50 mL) and the solids werefiltered off and washed with a small amount of CH₂Cl₂ and dried undervacuum at 50° C. to give2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-7-methyl-1H-indol-3-yl)ethanone6a (6.85 g) as an off-white solid.

Synthesis of Intermediate 6b

At 0° C., a solution of phenyltrimethylammonium tribromide [CAS4207-56-1] (8.2 g, 21.8 mmol) in THF (150 mL) was added dropwise to asolution of2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-7-methyl-1H-indol-3-yl)ethanone6a (6.8 g, 19.8 mmol) in THF (250 mL). The mixture was stirred at roomtemperature for 2 h.

The precipitate was filtered off and washed with THF. The filtrate wasconcentrated under reduced pressure. The residue was crystallized fromCH₂Cl₂. The precipitate was filtered off, wash with CH₂Cl₂ (2×) anddried under vacuum at 50° C. to give2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-7-methyl-1H-indol-3-yl)ethanone6b (5.38 g).

Synthesis of Compound 6 and Chiral Separation of Enantiomers 6A and 6B

A mixture of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-7-methyl-1H-indol-3-yl)ethanone6b (1.96 g, 4.65 mmol), 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (1.40 g, 6.97 mmol) and diisopropylethylamine (1.20 mL, 6.97mmol) in CH₃CN (50 mL) was heated overnight under reflux. The solventswere removed under reduced pressure. The residue was dissolved in CH₂Cl₂and washed with 0.5N HCl and water, dried over MgSO₄, filtered andevaporated under reduced pressure. The residue was purified by flashchromatography on silica gel (Stationary phase: Biotage® SNAP Ultra 100g, Mobile phase: EtOAc:EtOH(3:1)/heptane gradient 0/100 to 50/50). Thepure fractions were combined and evaporated under reduced pressure togive2-(4-chloro-2-methoxy-phenyl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)-1-(6-methoxy-7-methyl-1H-indol-3-yl)ethanone(Compound 6, 1.0 g) as a racemic mixture.

The chiral separation of the enantiomers of Compound 6 (1.0 g) wasperformed via Preparative Chiral SFC (Stationary phase: Chiralcel®Diacel OD 20×250 mm, Mobile phase: CO₂, EtOH containing 0.2% iPrNH₂).The product fractions were combined and evaporated. The first elutedenantiomer was solidified by trituration with a MeOH/water (1/1)mixture. The solids were filtered off and dried under vacuum at 50° C.to provide Enantiomer 6A (368 mg) as an amorphous white powder. Thesecond eluted enantiomer was solidified by trituration with a MeOH/water(1/1) mixture. The solids were filtered off and dried under vacuum at50° C. to provide Enantiomer 6B (303 mg) as an amorphous white powder.

Enantiomer 6A:

¹H NMR (360 MHz, DMSO-d₆) δ ppm 2.29 (s, 3H) 3.10 (s, 3H) 3.72 (s, 3H)3.80 (s, 3H) 4.02 (s, 3H) 6.24 (d, J=7.7 Hz, 1H) 6.56-6.59 (m, 1H)6.59-6.62 (m, 1H) 6.92 (t, J=1.6 Hz, 1H) 6.93-6.99 (m, 2H) 7.06 (d,J=7.7 Hz, 1H) 7.13 (d, J=1.8 Hz, 1H) 7.35 (d, J=8.4 Hz, 1H) 7.94 (d,J=8.4 Hz, 1H) 8.35 (s, 1H) 11.91 (br s, 1H)

LC/MS (method LC-A): R_(t) 1.18 min, MH⁺543

[α]_(D) ²⁰: +122.9° (c 0.48, DMF)

Chiral SFC (method SFC-E): R_(t) 4.15 min MH⁺543, chiral purity 100%.

Enantiomer 6B:

¹H NMR (360 MHz, DMSO-d₆) δ ppm 2.29 (s, 3H) 3.10 (s, 3H) 3.72 (s, 3H)3.80 (s, 3H) 4.02 (s, 3H) 6.24 (d, J=7.7 Hz, 1H) 6.57-6.59 (m, 1H)6.59-6.62 (m, 1H) 6.92 (t, J=1.8 Hz, 1H) 6.93-7.00 (m, 2H) 7.06 (d,J=7.7 Hz, 1H) 7.13 (d, J=1.8 Hz, 1H) 7.35 (d, J=8.1 Hz, 1H) 7.94 (d,J=8.8 Hz, 1H) 8.35 (d, J=2.2 Hz, 1H) 11.91 (br s, 1H)

LC/MS (method LC-A): R_(t) 1.22 min, MH⁺543

[α]_(D) ²⁰: −120.6° (c 0.2755, DMF)

Chiral SFC (method SFC-E): R_(t) 4.50 min, MH⁺543, chiral purity 99.35%.

Example 7: Synthesis of2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-5-methyl-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)ethanone(Compound 7) and Chiral Separation into Enantiomers 7A and 7B

Synthesis of Intermediate 7a

A solution of 6-fluoro-5-methyl-1H-indole [CAS 162100-95-0] (1.7 g, 11.4mmol) in CH₂Cl₂ (100 mL) was cooled to 0° C. under N₂-atmosphere. Asolution of diethylaluminum chloride 1M in hexane (17.1 mL, 17.1 mmol)was added dropwise and the resulting mixture was kept at 0° C. for 15min. A solution of 2-(4-chloro-2-methoxyphenyl)acetyl chloride 1a (3.50g, 16 mmol) in CH₂Cl₂ (50 mL) was added dropwise. Stirring was continuedat 0° C. for 1 h and at room temperature for 2 h. The reaction mixturewas poured out in a stirring ice/Rochelle salt solution.

After the ice had melted, the mixture was filtered over Dicalite® andthe filter cake was washed several times with THF. The filtrates werecombined. The layers were separated and the organic layer was washedwith brine, dried over MgSO₄, filtered and evaporated under reducedpressure. The solid residue was suspended in CH₂Cl₂ (30 mL), theprecipitate was filtered off and dried under vacuum at 50° C. to provide2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-5-methyl-1H-indol-3-yl)ethanone7a (2.76 g).

Synthesis of Intermediate 7b

A stirred solution of2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-5-methyl-1H-indol-3-yl)ethanone7a (2.76 g, 8.32 mmol) in THF (350 mL) was cooled to 0° C. A solution ofphenyltrimethylammonium tribromide [CAS 4207-56-1] (3.44 g, 9.15 mmol)in THF (50 mL) was added dropwise. The reaction mixture was stirred at0° C. for 2 h and at room temperature for 2 h. The solids were removedby filtration and washed with THF. The combined filtrates wereevaporated under reduced pressure. The residue was mixed with EtOAc (50mL). The solids were isolated by filtration, washed with a small amountof EtOAc and dried under vacuum at 50° C. to provide2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-5-methyl-1H-indol-3-yl)ethanone7b (3.21 g) as a white solid, which was used without furtherpurification in the next step.

Synthesis of Compound 7 and Chiral Separation of Enantiomers 7A and 7B

A mixture2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-fluoro-5-methyl-1H-indol-3-yl)ethanone7b (1.6 g, 3.90 mmol), 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (1.18 g, 5.84 mmol) and diisopropylethylamine (671 μL, 3.90mmol) in CH₃CN (100 mL) was stirred overnight at 85° C. The reactionmixture was concentrated under reduced pressure. The residue wasdissolved in CH₂Cl₂ (100 mL), washed with 1N HCl (100 mL) and water (100mL), dried over MgSO₄, filtered and evaporated under reduced pressure.The residue was purified by column chromatograph (Stationary phase:Grace Reveleris® silica 120 g, Mobile phase: EtOAc:EtOH(3:1)/heptanegradient 0/100 to 50/50). The desired fractions were combined andevaporated under reduced pressure. The residue was precipitated fromCH₂Cl₂/heptane. The solids were isolated by filtration and washed withCH₂Cl₂/heptane (1/1). The crude product was further purified byPreparative HPLC (Stationary phase: Uptisphere® C18 ODB—10 μm, 200 g, 5cm, Mobile phase: 0.25% NH₄HCO₃ solution in water, CH₃CN). The productfractions were combined and evaporated under reduced pressure. The solidresidue was mixed with EtOAc (20 mL) and the solids were isolated byfiltration and washed with a small amount of EtOAc to provide2-(4-chloro-2-methoxy-phenyl)-1-(6-fluoro-5-methyl-1H-indol-3-yl)-2-((3-methoxy-5-(methylsulfonyl)-phenyl)amino)ethanone(Compound 7, 341 mg) as a racemic mixture. The filtrate was evaporatedunder reduced pressure and the residue was taken up with MeOH. Afterstirring for 30 min, the solids were isolated by filtration to provide asecond crop of Compound 7 (92 mg).

The chiral separation of the enantiomers of Compound 7 (402 mg) wasperformed via Normal Phase Chiral separation (Stationary phase:(S,S)-Whelk-01, Mobile phase: 100% methanol). The product fractions werecombined and evaporated to provide Enantiomer 7A as the first elutedproduct and Enantiomer 7B as the second eluted product. Enantiomer 7Awas further purified by flash chromatography on silica gel (Stationaryphase: Grace Reveleris® silica 12 g, Mobile phase: heptane/EtOAc/EtOH100/0/0 to 40/45/15). The desired fractions were combined and evaporatedunder reduced pressure. The residue was triturated with H₂O (1.75 mL)and MeOH (0.75 mL). The solids were filtered off, washed (2×) withH₂O/MeOH 7/3, and dried under vacuum at 50° C. to provide Enantiomer 7A(48 mg). Enantiomer 7B was further purified by flash chromatography onsilica gel (Stationary phase: Grace Reveleris® silica 12 g, Mobilephase: heptane/EtOAc/EtOH 100/0/0 to 40/45/15). The desired fractionswere combined and evaporated under reduced pressure. The residue wastriturated with H₂O (1.75 mL) and MeOH (0.75 mL). The solids werefiltered off, washed (2×) with H₂O/MeOH 7/3, and dried under vacuum at50° C. to provide Enantiomer 7B (43 mg).

Compound 7:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 2.30 (d, J=0.9 Hz, 3H) 3.09 (s, 3H) 3.72(s, 3H) 4.00 (s, 3H) 6.22 (d, J=7.7 Hz, 1H) 6.54-6.63 (m, 2H) 6.92 (t,J=1.5 Hz, 1H) 6.97 (dd, J=8.3, 1.9 Hz, 1H) 7.01 (d, J=7.7 Hz, 1H) 7.12(d, J=1.8 Hz, 1H) 7.22 (d, J=10.2 Hz, 1H) 7.35 (d, J=8.4 Hz, 1H) 8.02(d, J=7.7 Hz, 1H) 8.37 (s, 1H) 11.97 (br s, 1H)

LC/MS (method LC-A): R_(t) 1.19 min, MH⁺531

Enantiomer 7A:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 2.30 (d, J=1.5 Hz, 3H) 3.09 (s, 3H) 3.72(s, 3H) 4.00 (s, 3H) 6.22 (d, J=7.9 Hz, 1H) 6.56-6.60 (m, 2H) 6.91 (t,J=1.7 Hz, 1H) 6.97 (dd, J=8.3, 2.1 Hz, 1H) 7.01 (d, J=7.7 Hz, 1H) 7.12(d, J=2.0 Hz, 1H) 7.22 (d, J=10.1 Hz, 1H) 7.34 (d, J=8.1 Hz, 1H) 8.02(d, J=7.7 Hz, 1H) 8.37 (s, 1H) 11.96 (s, 1H)

LC/MS (method LC-A): R_(t) 1.15 min, MH⁺531

[α]_(D) ²⁰: −163.2° (c 0.435, DMF)

Chiral SFC (method SFC-E): R_(t) 4.26 min, MH⁺531, chiral purity 100%.

Enantiomer 7B:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 2.30 (d, J=1.5 Hz, 3H) 3.09 (s, 3H) 3.72(s, 3H) 4.00 (s, 3H) 6.22 (d, J=7.7 Hz, 1H) 6.57-6.61 (m, 2H) 6.92 (t,J=1.8 Hz, 1H) 6.97 (dd, J=8.1, 2.0 Hz, 1H) 7.01 (d, J=7.7 Hz, 1H) 7.12(d, J=2.0 Hz, 1H) 7.22 (d, J=10.0 Hz, 1H) 7.35 (d, J=8.4 Hz, 1H) 8.02(d, J=7.9 Hz, 1H) 8.37 (d, J=2.4 Hz, 1H) 11.97 (s, 1H)

LC/MS (method LC-A): R_(t) 1.15 min, MH⁺531

[α]_(D) ²⁰: +166.6° (c 0.5, DMF)

Chiral SFC (method SFC-E): R_(t) 3.78 min, MH⁺531, chiral purity 100%.

Example 8: synthesis of2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methyl-sulfonyl)phenyl)amino)-1-(5-(trifluoromethyl)-1H-indol-3-yl)ethanone(Compound 8) and Chiral Separation into Enantiomers 8A and 8B

Synthesis of Intermediate 8a

At 0° C., under a N₂-flow, sodium hydride (2.48 g, 64.8 mmol) was addedportionwise to a mixture of 5-(trifluoromethyl)-1H-indole [CAS100846-24-0] (10 g, 54.0 mmol) in DMF (150 mL) and the reaction mixturewas stirred at 0° C. for 30 min. A solution of tosyl chloride (11.3 g,59.4 mmol) in DMF (50 mL) was added dropwise and the resulting mixturewas stirred at room temperature for 3 h. At 0° C., the mixture wasquenched by the addition of water. The precipitate was filtered off anddried overnight under vacuum at 70° C. to give1-tosyl-5-(trifluoromethyl)-1H-indole 8a (18.4 g).

Synthesis of Intermediate 8b

Titanium(IV) chloride (2.4 mL, 21.9 mmol) was added dropwise at roomtemperature to a solution of 1-tosyl-5-(trifluoromethyl)-1H-indole 8a(3.7 g, 10.95 mmol) and 2-(4-chloro-2-methoxyphenyl)acetyl chloride 1a(4.8 g, 21.9 mmol, synthesis: see Example 1) in 1,2-dichloroethane (120mL). The reaction was stirred at room temperature for 2 h. Ice-water wasadded. The reaction mixture was extracted with EtOAc. The organic layerwas dried over MgSO₄, filtered, and the solvent was concentrated underreduced pressure. The residue was purified by column chromatography onsilica gel (15-40 μm, 80 g, Mobile phase: CH₂Cl₂/MeOH 99.5/0.5). Thefractions containing Compound 8b were combined and the solvent wasevaporated under reduced pressure. The compound was taken up withCH₃CN/diisopropylether. The precipitate was filtered off and dried togive2-(4-chloro-2-methoxyphenyl)-1-(1-tosyl-5-(trifluoromethyl)-1H-indol-3-yl)ethanone8b (2.8 g).

Synthesis of Intermediate 8c

Lithium hydroxide (0.64 g, 15.3 mmol) was added to a solution of2-(4-chloro-2-methoxyphenyl)-1-(1-tosyl-5-(trifluoromethyl)-1H-indol-3-yl)ethanone8b (3.2 g, 6.13 mmol) in THF (18 mL) and water (6 mL). The mixture wasstirred at 30° C. for 1 h. Water and EtOAc were added. The organic layerwas separated, dried over MgSO₄, filtered, and the solvent wasevaporated under reduced pressure. The solid was taken up withdiisopropylether. The precipitate was filtered off and dried to give2-(4-chloro-2-methoxyphenyl)-1-(5-(trifluoromethyl)-1H-indol-3-yl)ethanone8c (2.1 g).

Synthesis of Intermediate 8d

At 0° C., a solution of phenyltrimethylammonium tribromide [CAS4207-56-1] (2.1 g, 5.7 mmol) in THF (60 mL) was added dropwise to amixture of2-(4-chloro-2-methoxyphenyl)-1-(5-(trifluoromethyl)-1H-indol-3-yl)ethanone8c (2.15 g, 5.7 mmol) in THF (60 mL). The mixture was stirred at 0° C.for 1 h and at room temperature for 4 h. The precipitate was filteredoff and washed with EtOAc. The combined filtrates were concentratedunder reduced pressure. The residue was dissolved in EtOAc. The organiclayer was washed with water, dried over MgSO₄, filtered and the solventwas evaporated under reduced pressure. The residue was taken up withdiisopropylether. The precipitate was filtered off and dried to give2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(5-(trifluoromethyl)-1H-indol-3-yl)-ethanone8d (2.5 g).

Synthesis of Compound 8 and Chiral Separation into Enantiomers 8A and 8B

A mixture of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(5-(trifluoromethyl)-1H-indol-3-yl)ethanone8d (1 g, 2.24 mmol), 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (496 mg, 2.46 mmol) and diisopropylethylamine (0.38 mL, 2.24mmol) in CH₃CN (50 mL) and THF (25 mL) was stirred at 70° C. for 24 h.The solution was concentrated under reduced pressure. The residue wasdissolved in EtOAc and the solution was washed with 1N HCl. The organiclayer was separated, dried over MgSO₄, filtered and the solvent wasevaporated under reduced pressure. The compound was crystallized fromdiisopropylether/CH₃CN to give2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)-1-(5-(trifluoro-methyl)-1H-indol-3-yl)ethanone(Compound 8, 310 mg) as a racemic mixture. The Enantiomers of Compound 8were separated via preparative Chiral SFC (Stationary phase: Chiralpak®AD-H 5 μm 250×20 mm, Mobile phase: 70% CO₂, 30% iPrOH+0.3% iPrNH₂) togive, after crystallization in petroleum ether/diisopropylether, 122 mgof the first eluted Enantiomer 8A and 128 mg of the second elutedEnantiomer 8B.

Compound 8:

¹H NMR (500 MHz, DMSO-d₆) δ ppm 3.10 (s, 3H) 3.72 (s, 3H) 3.99 (s, 3H)6.29 (d, J=7.9 Hz, 1H) 6.56-6.62 (m, 2H) 6.92 (s, 1H) 6.98 (dd, J=8.4,2.0 Hz, 1H) 7.09 (d, J=7.9 Hz, 1H) 7.13 (d, J=1.9 Hz, 1H) 7.36 (d, J=8.5Hz, 1H) 7.54 (dd, J=8.5, 1.6 Hz, 1H) 7.69 (d, J=8.5 Hz, 1H) 8.48 (s, 1H)8.61 (s, 1H) 12.45 (br s, 1H)

LC/MS (method LC-C): R_(t) 3.19 min, MH⁺567

Melting point: 168° C.

Enantiomer 8A:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H)6.29 (d, J=7.6 Hz, 1H) 6.60 (br s, 2H) 6.92 (s, 1H) 6.98 (dd, J=8.3, 1.8Hz, 1H) 7.07 (d, J=8.1 Hz, 1H) 7.13 (d, J=1.5 Hz, 1H) 7.36 (d, J=8.1 Hz,1H) 7.54 (d, J=8.1 Hz, 1H) 7.69 (d, J=8.6 Hz, 1H) 8.49 (s, 1H) 8.60 (s,1H) 12.41 (br s, 1H)

LC/MS (method LC-C): R_(t) 3.25 min, MH⁺567

[α]_(D) ²⁰: −119.2° (c 0.2727, DMF)

Chiral SFC (method SFC-F): R_(t) 2.64 min, MH⁺567, chiral purity 100%.

Enantiomer 8B:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H)6.29 (d, J=8.1 Hz, 1H) 6.60 (s, 2H) 6.92 (s, 1H) 6.98 (dd, J=8.6, 2.0Hz, 1H) 7.07 (d, J=8.1 Hz, 1H) 7.13 (d, J=2.0 Hz, 1H) 7.36 (d, J=8.6 Hz,1H) 7.54 (dd, J=8.6, 1.5 Hz, 1H) 7.69 (d, J=8.6 Hz, 1H) 8.49 (s, 1H)8.60 (s, 1H) 12.40 (br s, 1H)

LC/MS (method LC-C): R_(t) 3.25 min, MH⁺567

[α]_(D) ²⁰: +125.1° (c 0.2455, DMF)

Chiral SFC (method SFC-F): R_(t) 3.44 min, MH⁺567, chiral purity 100%.

Example 9: Synthesis of2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methyl-sulfonyl)phenyl)amino)-1-(5-(trifluoromethoxy)-1H-indol-3-yl)ethanone(Compound 9) and Chiral Separation into Enantiomers 9A and 9B

Synthesis of Intermediate 9a

A solution of 5-(trifluoromethoxy)-1H-indole [CAS 262593-63-5] (3 g,14.9 mmol) in CH₂Cl₂ (150 mL) was cooled to 0° C. under N₂-atmosphere. Asolution of diethylaluminum chloride 1M in hexane (22.4 mL, 22.4 mmol)was added dropwise and the resulting mixture was kept at 0° C. for 15min. A solution of 2-(4-chloro-2-methoxyphenyl)acetyl chloride 1a (4.57g, 20.9 mmol) in CH₂Cl₂ (100 mL) was added dropwise. Stirring wascontinued at 0° C. for 1 h and the reaction mixture was subsequentlystirred at room temperature for 4 h. The reaction mixture was poured outin a stirring ice/Rochelle salt solution. After the ice had melted, themixture was filtered over Dicalite® and the filter cake was washedseveral times with THF. The filtrates were combined. The layers wereseparated and the organic layer washed with brine, dried over MgSO₄,filtered and evaporated under reduced pressure. The residue wastriturated with CH₂Cl₂ (50 mL). The resulting precipitate was filteredoff and dried under vacuum at 50° C. to provide2-(4-chloro-2-methoxy-phenyl)-1-(5-(trifluoromethoxy)-1H-indol-3-yl)ethanone9a (4.39 g).

Synthesis of Intermediate 9b

A stirred solution of2-(4-chloro-2-methoxyphenyl)-1-(5-(trifluoromethoxy)-1H-indol-3-yl)ethanone9a (4.39 g, 11.4 mmol) in THF (200 mL) was cooled to 0° C. A solution ofphenyltrimethylammonium tribromide [CAS 4207-56-1] (4.73 g, 12.6 mmol)in THF (100 mL) was added dropwise. The resulting suspension was stirredat room temperature for 2 h. The solids were removed by filtration andwashed with THF. The combined filtrates were evaporated under reducedpressure. The residue was mixed with EtOAc (30 mL). The solids wereisolated by filtration, washed with a small amount of EtOAc and driedunder vacuum at 50° C. to provide2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(5-(trifluoromethoxy)-1H-indol-3-yl)ethanone9b (5.0 g) as a white solid, which was used without further purificationin the next step.

Synthesis of Compound 9 and Chiral Separation of Enantiomers 9A and 9B

A mixture2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(5-(trifluoromethoxy)-1H-indol-3-yl)ethanone9b (2.5 g, 5.40 mmol), 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (1.49 g, 7.38 mmol) and diisopropylethylamine (931 μL, 5.40mmol) in CH₃CN (100 mL) was stirred overnight at 90° C. The reactionmixture was concentrated under reduced pressure. The residue wasdissolved in CH₂Cl₂ (100 mL), washed with 1N HCl (100 mL) and water (100mL), dried over MgSO₄, filtered and evaporated under reduced pressure.The residue was purified by column chromatograph (Stationary phase:Grace Reveleris® silica 120 g, Mobile phase: EtOAc:EtOH(3:1)/heptanegradient 0/100 to 50/50). The desired fractions were combined andevaporated under reduced pressure. The residue was precipitated fromEtOAc (10 mL) while stirring. The solids were isolated by filtration andwashed with a small amount of EtOAc to provide2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)-1-(5-(trifluoro-methoxy)-1H-indol-3-yl)ethanone(Compound 9, 477 mg) as a racemic mixture. The filtrate was evaporatedunder reduced pressure and the residue was taken up with EtOAc (5 mL).After overnight stirring, the solids were isolated by filtration andwashed with EtOAc to provide a second crop of Compound 9 (216 mg).

The chiral separation of the enantiomers of Compound 9 (663 mg) wasperformed via Normal Phase Chiral separation (Stationary phase: AS 20μm, Mobile phase: 100% methanol). The product fractions were combinedand evaporated to provide Enantiomer 9A as the first eluted product andEnantiomer 9B as the second eluted product. Enantiomer 9A was stirred upin H₂O (2 mL) and MeOH (3 mL) at 40° C. The solids were filtered off,washed (3×) with H₂O/MeOH 1/1, and dried under vacuum at 45° C. toprovide Enantiomer 9A (151 mg). Enantiomer 9B was further purified byflash chromatography on silica gel (Stationary phase: Grace Reveleris®silica 12 g, Mobile phase: heptane/EtOAc/EtOH 100/0/0 to 40/45/15). Thedesired fractions were combined, evaporated under reduced pressure andco-evaporated with EtOAc. The residue was stirred up in MeOH (5 mL) andprecipitated by the slow addition of H₂O (4 mL). The solids werefiltered off, washed (3×) with H₂O/MeOH 1/1, and dried under vacuum at50° C. to provide Enantiomer 9B (132 mg).

Compound 9:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H)6.26 (d, J=7.9 Hz, 1H) 6.57-6.62 (m, 2H) 6.91 (t, J=1.9 Hz, 1H) 6.98(dd, J=8.4, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13 (d, J=2.0 Hz, 1H)7.22 (dd, J=8.6, 2.2 Hz, 1H) 7.36 (d, J=8.4 Hz, 1H) 7.59 (d, J=8.8 Hz,1H) 8.06 (d, J=0.9 Hz, 1H) 8.55 (s, 1H) 12.28 (br s, 1H)

LC/MS (method LC-A): R_(t) 1.31 min, MH⁺583

Enantiomer 9A:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H)6.26 (d, J=7.9 Hz, 1H) 6.55-6.62 (m, 2H) 6.91 (t, J=1.5 Hz, 1H) 6.98(dd, J=8.4, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13 (d, J=2.0 Hz, 1H)7.21 (dd, J=8.8, 1.8 Hz, 1H) 7.36 (d, J=8.4 Hz, 1H) 7.59 (d, J=8.8 Hz,1H) 8.07 (d, J=0.9 Hz, 1H) 8.55 (s, 1H) 12.29 (br s, 1H)

LC/MS (method LC-A): R_(t) 1.20 min, MH⁺583

[α]_(D) ²⁰: +130.3° (c 0.555, DMF)

Chiral SFC (method SFC-E): R_(t) 3.10 min, MH⁺583, chiral purity 100%.

Enantiomer 9B:

¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H)6.26 (d, J=7.9 Hz, 1H) 6.56-6.62 (m, 2H) 6.92 (t, J=2.0 Hz, 1H) 6.98(dd, J=8.1, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13 (d, J=2.0 Hz, 1H)7.22 (dd, J=8.8, 1.8 Hz, 1H) 7.36 (d, J=8.4 Hz, 1H) 7.59 (d, J=8.8 Hz,1H) 8.07 (d, J=0.9 Hz, 1H) 8.55 (s, 1H) 12.30 (br s, 1H)

LC/MS (method LC-A): R_(t) 1.20 min, MH⁺583

[α]_(D) ²⁰: −133.2° (c 0.5, DMF)

Chiral SFC (method SFC-E): R_(t) 3.50 min, MH⁺583, chiral purity 100%.

Example 10: Synthesis of2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)-1-(6-methoxy-5-(trifluoromethoxy)-1H-indol-3-yl)-ethanone(Compound 10) and Chiral Separation into Enantiomers 10A and 10B

Synthesis of Intermediate 10a

To a cooled (−15° C.) solution of3-methoxy-4-(trifluoromethoxy)benzaldehyde [CAS 853771-90-1] (50 g, 230mmol) and ethyl azidoacetate (89 g, 690 mmol) in EtOH (400 mL) was addeddropwise, over a period of 2 h, a solution of NaOEt (0.69 mol, preparedfrom 15.9 g of Na and 700 mL of EtOH). The reaction mixture was stirredat room temperature overnight. After cooling on an ice-bath, thereaction was quenched with a saturated NH₄Cl solution (1.2 L), andstirred for 10 min. The precipitate was filtered off, washed with water,and dried to give (Z)-ethyl2-azido-3-(3-methoxy-4-(trifluoromethoxy)phenyl)acrylate 10a (32 g) as ayellowish solid.

Synthesis of Intermediate 10b

A solution of (Z)-ethyl2-azido-3-(3-methoxy-4-(trifluoromethoxy)phenyl)acrylate 10a (3 g, 10mmol) in xylene (40 mL) was heated under reflux overnight. After coolingto room temperature, the solvent was evaporated to dryness. The residuewas triturated with hexane (50 mL) and the precipitate was filtered offto afford methyl 6-methoxy-5-(trifluoromethoxy)-1H-indole-2-carboxylate10b (yield: 1.4-1.6 g) as a yellow solid.

Synthesis of Intermediate 10c

To a mixture of methyl6-methoxy-5-(trifluoromethoxy)-1H-indole-2-carboxylate 10b (25 g, 87mmol) in MeOH/H₂O (2/1, 300 mL) was added NaOH (7 g, 175 mmol) and themixture was heated under reflux until a clear solution was obtained.After cooling to room temperature, most of the methanol was removedunder reduced pressure and the remaining aqueous solution was acidifiedwith conc. HCl to pH 3-4. The product was extracted with EtOAc (2×250mL). The combined organic layers were washed with brine, dried, andevaporated under reduced pressure to give6-methoxy-5-(trifluoromethoxy)-1H-indole-2-carboxylic acid 10c (22.7 g)as a grey solid.

Synthesis of Intermediate 10d

A suspension of 6-methoxy-5-(trifluoromethoxy)-1H-indole-2-carboxylicacid 10c (7.5 g, 27 mmol) and Cu (1.22 g, 0.7 equiv.) in quinoline (150mL) was heated to 220-230° C. under inert atmosphere for 12 h. Aftercooling to room temperature, the mixture was diluted with methyltert-butyl ether (MTBE, 400 mL) and washed with a saturated aqueousNaHSO₄ solution (2×500 mL). The organic layer was dried over MgSO₄,filtered through short pad of silica gel, and evaporated under reducedpressure. The residue was purified by column chromatography to afford6-methoxy-5-(trifluoromethoxy)-1H-indole 10d (3.75 g) as a yellow solid.

Synthesis of Intermediate 10e

A solution of 6-methoxy-5-(trifluoromethoxy)-1H-indole 10d (1.61 g, 6.96mmol) in CH₂Cl₂ (150 mL) was cooled to 0° C. under N₂-atmosphere. Asolution of diethylaluminum chloride 1M in hexane (10.4 mL, 10.4 mmol)was added dropwise and the resulting mixture was kept at 0° C. for 30min. A solution of 2-(4-chloro-2-methoxyphenyl)acetyl chloride 1a (2.28g, 10.4 mmol) in CH₂Cl₂ (75 mL) was added dropwise. Stirring wascontinued at 0° C. for 1 h and at room temperature for 1 h. The reactionmixture was cooled to 0° C. and a solution of potassium sodium tartratetetrahydrate (Rochelle salt, 3.93 g, 13.9 mmol) in water (6 mL) wasadded dropwise. The reaction mixture was stirred for 30 min at 0° C. THF(200 mL) was added and the reaction mixture was stirred at roomtemperature for 20 min. Na₂SO₄ (25 g) was added, the mixture was stirredovernight, filtered over Dicalite® and the filter cake was washedseveral times with THF (4×150 mL). The filtrates were combined andevaporated under reduced pressure. The solid residue was stirred up in amixture of diisopropyl ether (25 mL) and EtOAc (2 mL). The solids werefiltered off, washed with DIPE (3×) and dried under vacuum at 50° C. toprovide2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-5-(trifluoromethoxy)-1H-indol-3-yl)ethanone10e (3.6 g).

Synthesis of Intermediate 10f

A stirred solution of2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-5-(trifluoro-methoxy)-1H-indol-3-yl)ethanone10e (3.6 g, 6.53 mmol) in THF (130 mL) was cooled to 0° C., underN₂-atmosphere. Phenyltrimethylammonium tribromide [CAS 4207-56-1] (2.58g, 6.85 mmol) was added and the reaction mixture was stirred at 0° C.for 45 min and at room temperature for 1.5 h. The solids were removed byfiltration and washed with THF (2×). The combined filtrates wereevaporated under reduced pressure to provide2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-5-(trifluoromethoxy)-1H-indol-3-yl)ethanone10f (4.16 g), which was used without further purification in the nextstep.

Synthesis of Compound 10 and Chiral Separation of Enantiomers 10A and10B

A mixture2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(6-methoxy-5-(trifluoro-methoxy)-1H-indol-3-yl)ethanone10f (4.16 g, 6.50 mmol), 3-methoxy-5-(methyl-sulfonyl)aniline [CAS62606-02-4] (2.62 g, 13.0 mmol) and diisopropylethylamine (2.24 mL, 13.0mmol) in CH₃CN was stirred at room temperature for 2 days underN₂-atmosphere. Water (250 mL) was added and the product was extractedwith Et₂O (2×). The combined organic layers were dried over MgSO₄,filtered and evaporated under reduced pressure. The residue was purifiedby column chromatography (Stationary phase: Grace Reveleris® silica 100g, Mobile phase: heptane/EtOAc/EtOH gradient 100/0/0 to 40/45/15). Thedesired fractions were combined and evaporated under reduced pressure.The residue was further purified via preparative HPLC (Stationary phase:RP XBridge® Prep C18 OBD—10 μm, 50×150 mm, Mobile phase: 0.25% NH₄HCO₃solution in water, CH₃CN). The desired fractions were combined andevaporated under reduced pressure. The residue, containing racemic2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)-1-(6-methoxy-5-(trifluoromethoxy)-1H-indol-3-yl)ethanone(Compound 10, 380 mg), was submitted to chiral separation by preparativeSFC (Stationary phase: Chiralpak® Diacel AS 20×250 mm, Mobile phase:CO₂, EtOH+0.4% iPrNH₂). The product fractions were combined, evaporatedunder reduced pressure and co-evaporated with MeOH to provide Enantiomer10A as the first eluted product and Enantiomer 10B as the second elutedproduct. Both enantiomers were precipitated from a solvent mixture ofMeOH and water, filtered off and dried at 50° C. under vacuum to provideEnantiomer 10A (135 mg) and Enantiomer 10B (144 mg).

Enantiomer 10A:

¹H NMR (360 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.87 (s, 3H)3.99 (s, 3H) 6.22 (d, J=7.7 Hz, 1H) 6.55-6.59 (m, 2H) 6.88-6.91 (m, 1H)6.98 (dd, J=8.1, 1.8 Hz, 1H) 7.08 (d, J=7.7 Hz, 1H) 7.13 (d, J=2.2 Hz,1H) 7.21 (s, 1H) 7.34 (d, J=8.1 Hz, 1H) 8.02 (d, J=1.5 Hz, 1H) 8.41 (s,1H) 12.05 (br s, 1H) LC/MS (method LC-A): R_(t) 1.20 min, MH⁺613 [α]_(D)²⁰: +81.4° (c 0.29, DMF) Chiral SFC (method SFC-E): R_(t) 3.34 min,MH⁺613, chiral purity 100%.

Enantiomer 10B:

¹H NMR (360 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.87 (s, 3H)3.99 (s, 3H) 6.22 (d, J=7.7 Hz, 1H) 6.55-6.60 (m, 2H) 6.90 (t, J=1.6 Hz,1H) 6.98 (dd, J=8.2, 2.0 Hz, 1H) 7.08 (d, J=7.8 Hz, 1H) 7.13 (d, J=2.2Hz, 1H) 7.21 (s, 1H) 7.34 (d, J=8.4 Hz, 1H) 8.01 (d, J=1.1 Hz, 1H) 8.41(s, 1H) 12.08 (br s, 1H) LC/MS (method LC-A): R_(t) 1.20 min, MH⁺613

[α]_(D) ²⁰: −99.6° (c 0.261, DMF)

Chiral SFC (method SFC-E): R_(t) 3.69 min, MH⁺613, chiral purity 100%.

Example 11: Synthesis of2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methyl-sulfonyl)phenyl)amino)-1-(7-methyl-5-(trifluoromethoxy)-1H-indol-3-yl)ethanone(Compound 11) and Chiral Separation into Enantiomers 11A and 11B

Synthesis of Intermediate 11a

A mixture of boron(III) chloride 1M in CH₂Cl₂ (25.5 mL, 25.5 mmol) andaluminum(III) chloride (3.40 g, 25.5 mmol) was diluted with CH₂Cl₂ (20mL) and cooled on an ice-bath under N₂-atmosphere. A solution of2-methyl-4-(trifluoro-methoxy)aniline [CAS 86256-59-9] (4.88 g, 25.5mmol) and chloroacetonitrile (3.24 mL, 51.0 mmol) in CH₂Cl₂ (7.5 mL) wasadded dropwise. After addition, the ice-bath was removed and the mixturewas heated under reflux for 8 h. The mixture was cooled again to 0° C.using an ice-bath. 2N HCl (75 mL) was added dropwise, causing heavyprecipitation. The resulting suspension was heated under reflux for 90min, and cooled to room temperature. The solids were removed byfiltration. The filter cake was washed with CH₂Cl₂ (4×). The filtrateswere combined and the phases were separated. The organic layer wasisolated, washed with an aqueous NaHCO₃ solution, dried over MgSO₄,filtered and evaporated under reduced pressure. The residue was purifiedby flash chromatography (Stationary phase: Biotage® SNAP Ultra Silica100 g, Mobile phase: heptane/CH₂Cl₂ gradient 100/0 to 0/100). Thedesired fractions were combined and concentrated to a residual volume of30 mL. The precipitate was filtered off, washed with heptane and CH₂Cl₂,and dried under vacuum at 50° C. to provide1-(2-amino-3-methyl-5-(trifluoromethoxy)phenyl)-2-chloroethanone 11a(1.37 g). The filtrate was concentrated under reduced pressure. Thesolid residue was stirred up in a mixture of heptane (20 mL) anddiisopropyl ether (3 mL), filtered off, washed with heptane (3×) anddried under vacuum at 50° C. to provide a second fraction of 11a (0.24g).

Synthesis of Intermediate 11 b

Sodium borohydride (326 mg, 8.61 mmol) was added to a stirred solutionof 1-(2-amino-3-methyl-5-(trifluoromethoxy)phenyl)-2-chloroethanone 11a(1.92 g, 7.17 mmol) in tert-butanol (50 mL) and water (5 mL). Thereaction mixture was stirred at room temperature for 30 min and at 90°C. for 2.5 h. Water (50 mL) was added and the product was extracted withdiethyl ether (2×). The combined organic layers were washed with brine,dried over MgSO₄, filtered and evaporated under reduced pressure. Theresidue was purified by flash chromatography (Stationary phase: Biotage®SNAP Ultra Silica 25 g, Mobile phase: heptane/EtOAc gradient 100/0 to20/80). The desired fractions were combined, concentrated under reducedpressure, co-evaporated with heptane and dried under vacuum at 50° C. toprovide 7-methyl-5-(trifluoromethoxy)-1H-indole 11 b (1.2 g).

Synthesis of Intermediate 11c

A mechanically stirred solution of7-methyl-5-(trifluoromethoxy)-1H-indole 11 b (1.5 g, 6.97 mmol) inCH₂Cl₂ (100 mL) was cooled to 0° C. under N₂-atmosphere. A solution ofdiethylaluminum chloride 1M in hexane (10.5 mL, 10.5 mmol) was addeddropwise and the resulting mixture was kept at 0° C. for 25 min. Asolution of 2-(4-chloro-2-methoxyphenyl)acetyl chloride 1a (2.29 g, 10.5mmol) in CH₂Cl₂ (40 mL) was added dropwise while keeping the reactiontemperature below 6° C. Stirring was continued at 0° C. for 1 h and thereaction mixture was subsequently stirred at room temperature for 1 h.The reaction mixture was cooled to 0° C. and a solution of Rochelle salt[CAS 6100-16-9] (3.94 g, 13.9 mmol) in water (4 mL) was added dropwise.After stirring for 1 h, the reaction mixture was filtered over Dicalite®and the filter cake was washed with THF (5×100 mL). The combinedfiltrates were evaporated under reduced pressure. The residue solidifiedupon standing overnight. The solids were stirred up in CH₃CN (5 mL),filtered off, washed with CH₃CN (3×1.5 mL) and dried under vacuum at 50°C. to provide2-(4-chloro-2-methoxyphenyl)-1-(7-methyl-5-(trifluoromethoxy)-1H-indol-3-yl)-ethanone11c (1.9 g).

Synthesis of Intermediate 11d

A stirred solution2-(4-chloro-2-methoxyphenyl)-1-(7-methyl-5-(trifluoromethoxy)-1H-indol-3-yl)ethanone11c (2.13 g, 5.35 mmol) in THF (80 mL) was cooled to 0° C., underN₂-atmosphere. Phenyltrimethylammonium tribromide [CAS 4207-56-1] (2.11g, 5.62 mmol) was added and the reaction mixture was stirred at 0° C.for 40 min and at room temperature for 2 h. The solids were removed byfiltration and washed with THF (2×). The combined filtrates wereevaporated under reduced pressure to provide2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(7-methyl-5-(trifluoromethoxy)-1H-indol-3-yl)ethanone11d (3.45 g), which was used without further purification in the nextstep.

Synthesis of Compound 11 and Chiral Separation of Enantiomers 11A and11B

A mixture of2-bromo-2-(4-chloro-2-methoxyphenyl)-1-(7-methyl-5-(trifluoro-methoxy)-1H-indol-3-yl)ethanone11d (3.45 g, 6.87 mmol), 3-methoxy-5-(methylsulfonyl)aniline [CAS62606-02-4] (2.76 g, 13.7 mmol) and diisopropylethylamine (2.37 mL, 13.7mmol) in CH₃CN (60 mL) was stirred at room temperature for 2 days underN₂-atmosphere. Water (125 mL) was added and the product was extractedwith Et₂O (2×). The combined organic layers were washed with brine,dried over MgSO₄, filtered and evaporated under reduced pressure. Theresidue was purified via preparative HPLC (Stationary phase: RP XBridge®Prep C18 OBD—10 μm, 50×150 mm, Mobile phase: 0.25% NH₄HCO₃ solution inwater, CH₃CN). The fractions containing product were combined andevaporated under reduced pressure to provide racemic2-(4-chloro-2-methoxyphenyl)-2-((3-methoxy-5-(methylsulfonyl)phenyl)amino)-1-(7-methyl-5-(trifluoromethoxy)-1H-indol-3-yl)ethanone(Compound 11, 1.74 g). The chiral separation of the enantiomers ofCompound 11 (1.74 g) was performed via Preparative SFC (Stationaryphase: Chiralpak® Diacel AS 20×250 mm, Mobile phase: CO₂, EtOH+0.4%iPrNH₂). The product fractions were combined and evaporated underreduced pressure to provide Enantiomer 11A as the first eluted productand Enantiomer 11B as the second eluted product. Both enantiomers wereprecipitated from a solvent mixture of MeOH and water, filtered off anddried at 50° C. under vacuum to provide Enantiomer 11A (777 mg) andEnantiomer 11B (712 mg).

Enantiomer 11A:

¹H NMR (600 MHz, DMSO-d₆) δ ppm 2.50 (s, 3H) 3.09 (s, 3H) 3.72 (s, 3H)4.00 (s, 3H) 6.28 (d, J=7.8 Hz, 1H) 6.56-6.63 (m, 2H) 6.92 (br s, 1H)6.97 (dd, J=8.4, 1.9 Hz, 1H) 7.05 (br s, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13(d, J=1.9 Hz, 1H) 7.35 (d, J=8.4 Hz, 1H) 7.90 (br s, 1H) 8.53 (s, 1H)12.41 (br s, 1H)

LC/MS (method LC-A): R_(t) 1.26 min, MH⁺597

[α]_(D) ²⁰: +81.3° (c 0.3455, DMF)

Chiral SFC (method SFC-E): R_(t) 2.96 min, MH⁺597, chiral purity 100%.

Enantiomer 11B:

¹H NMR (600 MHz, DMSO-d₆) δ ppm 2.51 (s, 3H) 3.09 (s, 3H) 3.72 (s, 3H)4.00 (s, 3H) 6.28 (d, J=7.9 Hz, 1H) 6.58-6.60 (m, 2H) 6.92 (t, J=1.8 Hz,1H) 6.97 (dd, J=8.4, 1.9 Hz, 1H) 7.05 (br s, 1H) 7.06 (d, J=7.9 Hz, 1H)7.13 (d, J=2.1 Hz, 1H) 7.35 (d, J=8.2 Hz, 1H) 7.89 (br s, 1H) 8.53 (s,1H) 12.37 (br s, 1H)

LC/MS (method LC-A): R_(t) 1.26 min, MH⁺597

[α]_(D) ²⁰: −87.4° (c 0.342, DMF)

Chiral SFC (method SFC-E): R_(t) 3.44 min, MH⁺597, chiral purity 100%.

Antiviral Activity of the Compounds of the Invention

DENV-2 Antiviral Assay

The antiviral activity of all the compounds of the invention was testedagainst the DENV-2 16681 strain which was labeled with enhanced greenfluorescent protein (eGPF; Table 1). The culture medium consists ofminimal essential medium supplemented with 2% of heat-inactivated fetalcalf serum, 0.04% gentamycin (50 mg/mL) and 2 mM of L-glutamine. Verocells, obtained from ECACC, were suspended in culture medium and 25 μLwas added to 384-well plates (2500 cells/well), which already containthe antiviral compounds. Typically, these plates contain a 5-fold serialdilution of 9 dilution steps of the test compound at 200 times the finalconcentration in 100% DMSO (200 nL). In addition, each compoundconcentration is tested in quadruplicate (final concentration range: 25μM-0.000064 μM or 2.5 μM-0.0000064 μM for the most active compounds).Finally, each plate contains wells which are assigned as virus controls(containing cells and virus in the absence of compound), cell controls(containing cells in the absence of virus and compound) and mediumcontrols (containing medium in the absence of cells, virus andcompounds). To the wells assigned as medium control, 25 μL of culturemedium was added instead of Vero cells. Once the cells were added to theplates, the plates were incubated for 30 minutes at room temperature toallow the cells to distribute evenly within the wells. Next, the plateswere incubated in a fully humidified incubator (37° C., 5% CO₂) untilthe next day. Then, DENV-2 strain 16681, labeled with eGFP, was added ata multiplicity of infection (MOI) of 0.5. Therefore, 15 μL of virussuspension was added to all the wells containing test compound and tothe wells assigned as virus control. In parallel, 15 μL of culturemedium was added to the medium and cell controls. Next, the plates wereincubated for 3 days in a fully humidified incubator (37° C., 5% CO₂).At the day of the read out, the eGFP fluorescence was measured using anautomated fluorescence microscope at 488 nm (blue laser). Using anin-house LIMS system, inhibition dose response curves for each compoundwere calculated and the half maximal effective concentration (EC₅₀) wasdetermined. Therefore, the percent inhibition (I) for every testconcentration is calculated using the following formula:I=100*(S_(T)−S_(CC))/(S_(VC)−S_(CC)); S_(T), S_(CC) and S_(VC) are theamount of eGFP signal in the test compound, cell control and viruscontrol wells, respectively. The EC₅₀ represents the concentration of acompound at which the virus replication is inhibited with 50%, asmeasured by a 50% reduction of the eGFP fluorescent intensity comparedto the virus control. The EC₅₀ is calculated using linear interpolation.

In parallel, the toxicity of the compounds was assessed on the sameplates. Once the read-out for the eGFP signal was done, 40 μL ofATPlite, a cell viability stain, was added to all wells of the 384-wellplates. ATP is present in all metabolically active cells and theconcentration declines very rapidly when the cells undergo necrosis orapoptosis. The ATPLite assay system is based on the production of lightcaused by the reaction of ATP with added luciferase and D-luciferin. Theplates were incubated for 10 minutes at room temperature. Next, theplates were measured on a ViewLux. The half maximal cytotoxicconcentration (CC₅₀) was also determined, defined as the concentrationrequired to reduce the luminescent signal by 50% compared to that of thecell control wells. Finally, the selectivity index (SI) was determinedfor the compounds, which was calculated as followed:SI=CC₅₀/EC₅₀.

TABLE 1 EC₅₀, CC₅₀, and SI for the compounds of the invention in theDENV-2 antiviral assay compound# EC₅₀ (μM) N CC₅₀ (μM) N SI N  1 0.000525 5.5 4 11500 4  1A 0.00026 8 4.3 8 19700 8  1B 0.012 6 6.5 6 530 6  20.00060 4 5.0 4 8410 4  2A 0.00026 4 4.8 4 22000 4  2B 0.026 4 7.4 4 2854  3 0.00058 4 >11 6 37700 4  3A 0.00025 5 7.2 5 29800 5  3B 0.00383 >9.7 5 2480 3  4 0.00039 4 5.9 4 14900 4  4A 0.00027 11 4.2 13 1690011  4B 0.036 5 12 5 341 5  5 0.00062 4 5.5 4 8780 4  5A 0.00041 5 5.0 512900 5  5B 0.068 4 13 4 206 4  6A 0.000068 8 >25 8 >65500 8  6B 0.019 411 4 603 4  7 0.00047 4 3.2 3 >7040 3  7A 0.013 3 6.8 3 538 3  7B0.00020 5 3.2 5 18500 5  8 0.00013 6 2.9 7 30400 6  8A 0.0030 3 7.4 32510 3  8B 0.000069 5 3.4 5 >40900 5  9 0.000074 6 3.1 8 >39100 6  9A0.000067 9 2.9 9 >37500 9  9B 0.0038 5 6.2 6 1480 5 10A 0.00012 3 2.6 322600 3 10B 0.0039 3 9.8 3 2530 3 11A 0.000085 3 2.6 3 30100 3 11B0.0041 3 9.2 3 2220 3 N = the number of independent experiments in whichthe compounds were tested.

Tetravalent reverse transcriptase quantitative-PCR (RT-qPCR) assay:Protocol A.

The antiviral activity of the compounds of the invention was testedagainst DENV-1 strain TC974#666 (NCPV; Table 6), DENV-2 strain 16681(Table 7), DENV-3 strain H87 (NCPV; Table 8) and DENV-4 strains H241(NCPV; Table 9A) and SG/06K2270DK1/2005 (Eden; Table 9B) in a RT-qPCRassay. Therefore, Vero cells were infected with either DENV-1, or -2, or-3, or -4 in the presence or absence of test compounds. At day 3post-infection, the cells were lysed and cell lysates were used toprepare cDNA of both a viral target (the 3′UTR of DENV; Table 2) and acellular reference gene ((3-actin, Table 2). Subsequently, a duplex realtime PCR was performed on a Lightcycler480 instrument. The generated Cpvalue is inversely proportional to the amount of RNA expression of thesetargets. Inhibition of DENV replication by test compound results in ashift of Cp's for the 3′UTR gene. On the other hand, if a test compoundis toxic to the cells, a similar effect on (3-actin expression will beobserved. The comparative ΔΔCp method is used to calculate EC₅₀, whichis based on the relative gene expression of the target gene (3′UTR)normalized with the cellular housekeeping gene ((3-actin).

TABLE 2 Primers and probes used for the real-time, quantitative RT-PCR.Primer/ probe Target Sequence^(a, b) F3utr258 DENV 5′-CGGTTAGAGG 3′-UTRAGACCCCTC-3′ R3utr425 DENV 5′-GAGACAGCAG 3′-UTR GATCTCTGGTC-3′ P3utr343DENV

-5′-AAGGACTAG-ZEN- 3′-UTR AGGTTAGAGGAGACCCCCC-3′-

Factin743 β-actin 5′-GGCCAGGTCATCACCATT-3′ Ractin876 β-actin5′-ATGTCCACGTCACACTTCATG-3′ Pactin773 β-actin

-5′-TTCCGCTGC-

- CCTGAGGCTCTC-3′-

^(a)Reporter dyes (FAM, HEX) and quenchers (ZEN and IABkFQ) elements areindicated in bold and italics. ^(b)The nucleotide sequence of theprimers and probes were selected from the conserved region in the 3′UTRregion of the dengue virus genome, based on the alignment of 300nucleotide sequences of the four dengue serotypes deposited in Genbank(Gong et al., 2013, Methods Mol Biol, Chapter 16).

The culture medium consisted of minimal essential medium supplementedwith 2% of heat-inactivated fetal calf serum, 0.04% gentamycin (50mg/mL) and 2 mM of L-glutamine. Vero cells, obtained from ECACC, weresuspended in culture medium and 75 μL/well was added in 96-well plates(10000 cells/well), which already contain the antiviral compounds.Typically, these plates contain a 5-fold serial dilution of 9 dilutionsteps of the test compound at 200 times the final concentration in 100%DMSO (500 nL; final concentration range: 25 μM-0.000064 μM or 2.5μM-0.0000064 μM for the most active compounds). In addition, each platecontains wells which are assigned as virus controls (containing cellsand virus in the absence of compound) and cell controls (containingcells in the absence of virus and compound). Once the cells were addedin the plates, the plates were incubated in a fully humidified incubator(37° C., 5% CO₂) until the next day. Dengue viruses serotype-1,2, 3 and4 were diluted in order to obtain a Cp of ˜22-24 in the assay.Therefore, 25 μL of virus suspension was added to all the wellscontaining test compound and to the wells assigned as virus control. Inparallel, 25 μL of culture medium was added to the cell controls. Next,the plates were incubated for 3 days in a fully humidified incubator(37° C., 5% CO₂). After 3 days, the supernatant was removed from thewells and the cells were washed twice with ice-cold PBS (˜100 μL). Thecell pellets within the 96-well plates were stored at −80° C. for atleast 1 day. Next, RNA was extracted using the Cells-to-CT™ lysis kit,according to the manufacturer's guideline (Life Technologies). The celllysates can be stored at −80° C. or immediately used in the reversetranscription step.

In preparation of the reverse transcription step, mix A (table 3A) wasprepared and 7.57 μL/well was dispensed in a 96-well plate. Afteraddition of 5 μL of the cell lysates, a five minute denaturation step at75° C. was performed (table 3B). Afterwards, 7.43 μL of mix B was added(table 3C) and the reverse transcription step was initiated (table 3D)to generate cDNA.

Finally, a RT-qPCR mix was prepared, mix C (table 4A), and 22.02 μL/wellwas dispensed in 96-well LightCycler qPCR plates to which 3 μL of cDNAwas added and the qPCR was performed according to the conditions intable 4B on a LightCycler 480.

Using the LightCycler software and an in-house LIMS system, doseresponse curves for each compound were calculated and the half maximaleffective concentration (EC₅₀) and the half maximal cytotoxicconcentration (CC₅₀) were determined.

TABLE 3 cDNA synthesis using Mix A, denaturation, Mix B and reversetranscription. A Mix A Plates 8 Sample 828 Reaction Vol. (μl) 20 Volumefor (μl) Concentration 1 x Mix Item Unit Stock Final sample samplesMilli-Q H₂O 7.27 6019.56 R3utr425 μM 20 0.27 0.15 124.20 Ractin876 μM 200.27 0.15 124.20 Volume mix/well (μl) 7.57 Cell lysates 5.00 BDenaturation step: Step Temp Time Denaturation 75° C. 5′ Hold  4° C.hold C Mix B Samples 864 Volume for (μl) Concentration 1 x Mix Item UnitStock Final sample samples Expand HIFI buffer 2 X 10.00 1.00 2.00 1728.0MgCl₂ mM 25.00 3.50 2.80 2419.2 dNTPs mM 10.00 1.00 2.00 1728.0 Rnaseinhibitor U/μl 40.00 1.00 0.50 432.0 Expand RT U/μl 50.00 0.33 0.13112.3 Total Volume Mix (μl) 7.43 D Protocol cDNA synthesis Step TempTime Rev transc 42° C. 30′ Denaturation 99° C.  5′ Hold  4° C. hold

TABLE 4 qPCR mix and protocol. A Mix C Samples 833 Reaction Vol. (μl) 25Volume for (μl) Concentration 1 x Mix Item Unit Stock Final samplesamples H₂O PCR grade 7.74 6447.42 Roche Roche 2xMM mix X 2 1 12.5010412.50 F3utr258 μM 20 0.3 0.38 316.54 R3utr425 μM 20 0.3 0.38 316.54P3utr343 μM 20 0.1 0.13 108.29 Factin743 μM 20 0.3 0.38 316.54 Ractin876μM 20 0.3 0.38 316.54 Pactin773 μM 20 0.1 0.13 108.29 Volume Mix/Tube(μl) 22.02 cDNA 3.00 B Protocol qPCR3 Step Temp Time Ramp ratepreincub/denat 95° C. 10 min 4.4 Denaturation 95° C. 10 sec 4.4 40cycles annealing 58° C. 1 min 2.2 Elongation 72° C. 1 sec 4.4 Cooling40° C. 10 sec 1.5

Tetravalent quantitative reverse transcriptase-PCR (RT-qPCR) assay:Protocol B.

The antiviral activity of the compounds of the invention was testedagainst DENV-1 strain Djibouti strain (D1/H/IMTSSA/98/606; Table 6),DENV-2 strain NGC (Table 7), DENV-3 strain H87 (Table 8) and DENV-4strain SG/06K2270DK1/2005 (Table 9B) in a RT-qPCR assay. Vero-B orVero-M cells (5×10⁴) were seeded in 96-well plates. One day later,culture medium was replaced with 100 μL assay medium containing a 2×, 3×or 5× serial dilution of the compound (concentration range: 50μg/mL-0.00038 μg/mL, 50 μg/mL-0.0076 μg/mL, and 50 μg/mL-0.00013 μg/mL,respectively) and 100 μL of dengue virus inoculum (DENV). Following a 2hour incubation period, the cell monolayer was washed 3 times with assaymedium to remove residual, non-adsorbed virus and cultures were furtherincubated for either 4 days (DENV-2 NGC) or 7 days (DENV-1 Djiboutistrain D1/H/IMTSSA/98/606, DENV-3 strain H87 prototype, DENV-4 strainH241, and DENV-4 strain EDEN) in the presence of the inhibitor.Supernatant was harvested and viral RNA load was determined by real-timequantitative RT-PCR. The 50% effective concentration (EC₅₀), which isdefined as the compound concentration that is required to inhibit viralRNA replication by 50%, was determined using logarithmic interpolation.

RNA was isolated from 100 μL (or in some circumstances 150 μL)supernatant with the NucleoSpin 96 Virus kit (Filter Service, Duren,Germany) as described by the manufacturer. The sequences of the TaqManprimers (DENV-For, DENV-Rev; Table 5) and TaqMan probes (DENV-ProbeTable 5) were selected from non-structural gene 3 (NS3) or NS5, of therespective flaviviruses using Primer Express software (version 2.0;Applied Biosystems, Lennik, Belgium). The TaqMan probe was fluorescentlylabelled with 6-carboxyfluorescein (FAM) at the 5′ end as the reporterdye, and with minor groove binder (MGB) at the 3′ end as the quencher(Table 5). One-step, quantitative RT-PCR was performed in a total volumeof 25 μL, containing 13.9375 μL H₂O, 6.25 μL master mix (Eurogentec,Seraing, Belgium), 0.375 μL forward primer, 0.375 μL reverse primer, 1μL probe, 0.0625 μL reverse transcriptase (Eurogentec) and 3 μL sample.RT-PCR was performed using the ABI 7500 Fast Real-Time PCR System(Applied Biosystems, Branchburg, New Jersey, USA) using the followingconditions: 30 min at 48° C. and 10 min at 95° C., followed by 40 cyclesof 15 s at 95° C. and 1 min at 60° C. The data was analyzed using theABI PRISM 7500 SDS software (version 1.3.1; Applied Biosystems). Forabsolute quantification, standard curves were generated using 10-folddilutions of template preparations of known concentrations.

TABLE 5 Primers and probes used for real-time, quantitative RT-PCR.Primer/ Sequence Probe (5′→3′)^(a) Source^(b) Target DENV-ForTCGGAGCCGGA DENV 2 NS3 GTTTACAAA NGC (SEQ ID N. 1) DENV-Rev TCTTAACGTCCGCCCATGAT (SEQ ID N. 2) DENV-Probe

-ATTCCACACA ATGTGGCAT-

(SEQ ID N. 3) DenS GGATAGACCAGAGAT DENV-1,  NS5 CCTGCTGT -3, -4(SEQ ID N. 4) DenAS1-3 CATTCCATTTT DENV-1, CTGGCGTTC -3 (SEQ ID N. 5)DenAS4 CAATCCATCTT DENV-4 GCGGCGCTC (SEQ ID N. 6) DEN_1-3

-CAGCATCA DENV-1, probe TTCCAGGCA -3 CAG-

(SEQ ID N. 7) DEN _4

-CAACATCA DENV-4 probe ATCCAGGCA CAG-

(SEQ ID N. 8) ^(a)Reporter dye (FAM) and quencher (MGB/TAMRA) elementsare indicated in bold and italics. ^(b)The nucleotide sequence andposition of the primers and probes within the genome were deduced fromthe nucleotide sequence of DENV 2 NGC (GenBank accession no. M29095;Irie et al., 1989), dengue virus serotype 1 Djibouti strainD1/H/IMTSSA/98/606 (Genbank Accession Number AF298808), dengue virusserotype 3 strain H87 prototype (c93130), dengue virus serotype 4 strainH241 (no sequences available), dengue virus serotype 4 strain EDEN (nosequences available)

Cytotoxic Assay

Potential cytotoxic effects of the compounds were evaluated inuninfected quiescent Vero-B or Vero-M cells. Cells were seeded at 5×10⁴cells/well in a 96-well plate in the presence of two-, three- orfive-fold serial dilutions (ranging from 50 μg/mL-0.0038 μg/mL, 50μg/mL-0.0076 μg/mL, and 50 μg/mL-0.00013 μg/mL, respectively) ofcompound and incubated for 4 to 7 days. Culture medium was discarded and100 μL3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxy-methoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium/phenazinemethosulfate(MTS/PMS; Promega, Leiden, The Netherlands) in PBS was added to eachwell. Following a 2-hour incubation period at 37° C., the opticaldensity was determined at 498 nm. Cytotoxic activity was calculatedusing the following formula: % cellviability=100×(OD_(compound)/OD_(CC)), where OD_(compound) and OD_(CC)correspond to the optical density at 498 nm of the uninfected cellcultures treated with compound and that of uninfected, untreated cellcultures, respectively. The 50% cytotoxic concentration (i.e., theconcentration that reduces the total cell number with 50%; CC₅₀) wascalculated using linear interpolation.

TABLE 6 EC₅₀, CC₅₀, and SI for the compounds against serotype 1 in theRT-qPCR assays Protocol A Protocol B RT-qPCR serotype 1 TC974#666RT-qPCR serotype 1 Djibouti compound# EC₅₀ (μM) N CC₅₀ (μM) N SI N EC₅₀(μM) N CC₅₀ (μM) N SI N  1A 0.0025 9 5.0 9 1950 9 <0.015 3 8 6 >533 3 2A 0.0024 6 5.3 6 2190 6 <0.014 2 7.3 3 >523 2  3A 0.0042 6 5.5 5 13605 ND ND ND ND ND ND  4A 0.00097 8 5.1 8 4400 8 <0.014 3 4.3 4 >306 3  5A0.0036 6 5.2 6 1460 6 <0.014 2 9.2 2 >658 2  6A 0.0016 6 >10 6 >8160 6<0.014 2 >92 3 >6571 2  7B 0.00040 3 2.2 2 6600 2 ND ND ND ND ND ND  8B0.00045 5 1.9 6 3130 4 ND ND ND ND ND ND  9A 0.00011 4 1.7 5 13300 4 NDND ND ND ND ND 10A 0.00027 2 1.6 2 5670 2 ND ND ND ND ND ND 11A 0.000132 >2.5 2 >22100 2 ND ND ND ND ND ND N = the number of independentexperiments in which the compounds were tested. ND: not determined.

TABLE 7 EC₅₀, CC₅₀, and SI for the compounds against serotype 2 in theRT-qPCR assays Protocol A Protocol B RT-qPCR serotype 2 16681 RT-qPCRserotype 2 NGC-Tongalike compound# EC50 (μM) N CC50 (μM) N SI N EC50(μM) N CC50 (μM) N SI N  1A 0.00028 7 3.8 12 15300 8 <0.00027 4 114 >40470 4  2A 0.00024 5 4.9 6 21500 5 <0.00024 1 11 1 >45833 1  3A0.00030 6 5.0 6 9970 6 ND ND ND ND ND ND  4A 0.00020 7 3.9 10 25400 60.00032 1 6.6 1 20339 1  5A 0.00034 5 5.8 6 19000 5 <0.00023 1 ND ND NDND  6A 0.00011 7 >10 6 >142306 6 <0.00024 1 >92 1 >383333 1  7B 0.000173 2.9 5 23600 3 ND ND ND ND ND ND  8B 0.00031 4 2.2 6 23400 4 ND ND NDND ND ND  9A 0.000057 3 2.2 4 31700 3 ND ND ND ND ND ND 10A 0.000057 31.6 3 28200 3 ND ND ND ND ND ND 11A 0.000051 3 >2.5 3 >69000 3 ND ND NDND ND ND N = the number of independent experiments in which thecompounds were tested. ND: not determined.

TABLE 8 EC₅₀, CC₅₀, and SI for the compounds against serotype 3 in theRT-qPCR assays Protocol A Protocol B RT-qPCR serotype 3 H87 RT-qPCRserotype 3 H87 compound# EC50 (μM) N CC50 (μM) N SI N EC50 (μM) N CC50(μM) N SI N  1A 0.023 7 3.7 5 169 5 <0.015 3 8.0 6 >533 3  2A 0.019 44.3 3 224 3 <0.014 1 7.3 3 >521 1  3A 0.048 4 4.1 3 67 3 ND ND ND ND NDND  4A 0.015 6 3.1 4 195 4 <0.014 1 4.3 4 >307 1  5A 0.053 4 4.4 2 75 20.022 1 9.2 2 422 1  6A 0.019 4 6.7 3 318 3 <0.014 1 >92 3 >6571 1  7B0.0078 3 1.6 3 240 3 ND ND ND ND ND ND  8B 0.0058 4 2.1 3 609 3 ND ND NDND ND ND  9A 0.0021 3 1.6 1 474 1 ND ND ND ND ND ND 10A 0.0037 3 1.0 3280 3 ND ND ND ND ND ND 11A 0.0012 3 >2.5 3 >2630 3 ND ND ND ND ND ND N= the number of independent experiments in which the compounds weretested. ND: not determined.

TABLE 9 EC₅₀, CC₅₀, and SI for the compounds against serotype 4 in theRT-gPCR assays A Protocol A RT-qPCR serotyoe 4 H241 EC50 CC50 compound#(μM) N (μM) N SI N 1A 0.093 10 3.0 9 30 9 2A 0.083 6 3.7 6 42 6 3A 0.116 3.8 4 37 4 4A 0.053 11 2.5 11 54 11 5A 0.10 6 4.0 6 39 6 6A 0.095 77.7 5 69 5 7B 0.044 5 2.2 5 53 5 8B 0.015 5 1.7 3 122 3 9A 0.012 5 1.5 5121 5 10A  0.011 3 1.6 2 127 2 11A  0.011 3 3.1 3 >250 3 B Protocol ART-qPCR serotype 4 EDEN EC₅₀ CC₅₀ compound# (μM) N (μM) N SI N 1A 0.00245 4.6 5 1927 5 2A 0.0013 2 5.0 2 3913 2 3A 0.0030 2 5.4 2 1802 2 4A0.00055 2 >2.5 1 >4520 1 5A 0.0029 2 5.5 2 1878 2 6A 0.00042 2 >102 >24085 2 N = the number of independent experiments in which thecompounds were tested.

The invention claimed is:
 1. A method of inhibiting Dengue viralreplication in an animal cell, comprising administering to the animalcell a compound of formula (I):

wherein R₁, R₂ and R₃ are selected from the group consisting of: R₁ isH, R₂ is F and R₃ is H or CH₃, R₁ is H, CH₃ or F, R₂ is OCH₃ and R₃ isH, R₁ is H, R₂ is OCH₃ and R₃ is CH₃, R₁ is CH₃, R₂ is F and R₃ is H, R₁is CF₃ or OCF₃, R₂ is H and R₃ is H, R₁ is OCF₃, R₂ is OCH₃ and R₃ is H,and R₁ is OCF₃, R₂ is H and R₃ is CH₃, or a stereoisomer,pharmaceutically acceptable salt, solvate or polymorph thereof.
 2. Themethod of claim 1, wherein the animal cell is a mammalian cell.
 3. Themethod of claim 1, wherein the animal cell is a human cell.
 4. Themethod of claim 1, wherein the compound is administered to the cellprior to being infected with Dengue virus.
 5. The method of claim 1,further comprising administering another antiviral agent to the cell. 6.The method of claim 1, wherein said compound is selected from the groupconsisting of:

or a stereoisomer, pharmaceutically acceptable salt, solvate orpolymorph thereof.
 7. The method of claim 1, wherein said compound isselected from the group consisting of: Enantiomer 1A wherein ¹H NMR (500MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 4.00 (s, 3H) 6.24 (d,J=7.9 Hz, 1H) 6.59 (s, 2H) 6.91 (s, 1H) 6.97 (dd, J=8.8, 2.2 Hz, 1H)7.02-7.10 (m, 2H) 7.12 (d, J=2.2 Hz, 1H) 7.27 (dd, J=9.6, 2.2 Hz, 1H)7.35 (d, J=8.2 Hz, 1H) 8.14 (dd, J=8.8, 5.7 Hz, 1H) 8.44 (s, 1H) 12.10(br. s., 1H) LC/MS (method LC-C): R_(t) 3.09 min, MH⁺ 517 [α]_(D) ²⁰:+130.3° (c 0.277, DMF) Chiral SFC (method SFC-D): R_(t) 3.41 min, MH⁺517, chiral purity 100%, Enantiomer 1B wherein ¹H NMR (400 MHz, DMSO-d₆)δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 4.00 (s, 3H) 6.24 (d, J=7.6 Hz, 1H)6.53-6.65 (m, 2H) 6.91 (s, 1H) 6.97 (dd, J=8.6, 2.0 Hz, 1H) 7.01-7.09(m, 2H) 7.12 (d, J=2.0 Hz, 1H) 7.27 (dd, J=9.6, 2.0 Hz, 1H) 7.35 (d,J=8.1 Hz, 1H) 8.14 (dd, J=8.6, 5.6 Hz, 1H) 8.43 (s, 1H) 12.09 (br. s.,1H) LC/MS (method LC-C): R_(t) 3.09 min, MH⁺ 517 [α]_(D) ²⁰: −135.3° (c0.283, DMF) Chiral SFC (method SFC-D): R_(t) 4.89 min, MH⁺ 517, chiralpurity 99.35%, Enantiomer 2A wherein ¹H NMR (500 MHz, DMSO-d₆) δ ppm2.37-2.39 (m, 3H) 3.09 (s, 3H) 3.72 (s, 3H) 4.01 (s, 3H) 6.26 (d, J=7.9Hz, 1H) 6.54-6.63 (m, 2H) 6.92 (s, 1H) 6.97 (dd, J=8.4, 1.9 Hz, 1H) 7.02(dd, J=9.9, 9.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13 (d, J=1.9 Hz, 1H)7.35 (d, J=8.4 Hz, 1H) 7.96 (dd, J=8.5, 5.4 Hz, 1H) 8.45 (s, 1H) 12.24(br. s., 1H) LC/MS (method LC-C): R_(t) 3.20 min, MH⁺ 531 [α]_(D) ²⁰:+104.5° (c 0.2545, DMF) Chiral SFC (method SFC-A): R_(t) 4.22 min, MH⁺531, chiral purity 100%, Enantiomer 2B wherein ¹H NMR (500 MHz, DMSO-d₆)δ ppm 2.36-2.41 (m, 3H) 3.09 (s, 3H) 3.72 (s, 3H) 4.01 (s, 3H) 6.26 (d,J=7.9 Hz, 1H) 6.57-6.64 (m, 2H) 6.92 (s, 1H) 6.97 (dd, J=8.2, 1.9 Hz,1H) 6.99-7.04 (m, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13 (d, J=1.9 Hz, 1H) 7.35(d, J=8.2 Hz, 1H) 7.96 (dd, J=8.7, 5.2 Hz, 1H) 8.45 (s, 1H) 12.24 (br.s., 1H) LC/MS (method LC-C): R_(t) 3.20 min, MH⁺ 531 [α]_(D) ²⁰: −104.1°(c 0.2536, DMF) Chiral SFC (method SFC-A): R_(t) 5.12 min, MH⁺ 531,chiral purity 99.53%, Enantiomer 3A wherein ¹H NMR (360 MHz, DMSO-d₆) δppm 3.09 (s, 3H) 3.72 (s, 3H) 3.77 (s, 3H) 4.01 (s, 3H) 6.22 (d, J=8.1Hz, 1H) 6.55-6.61 (m, 2H) 6.84 (dd, J=8.8, 2.2 Hz, 1H) 6.91 (t, J=1.8Hz, 1H) 6.94-7.00 (m, 2H) 7.07 (d, J=7.0 Hz, 1H) 7.13 (d, J=1.8 Hz, 1H)7.35 (d, J=8.4 Hz, 1H) 8.02 (d, J=8.8 Hz, 1H) 8.32 (d, J=2.9 Hz, 1H)11.87 (d, J=2.6 Hz, 1H) LC/MS (method LC-A): R_(t) 1.08 min, MH⁺ 529[α]_(D) ²⁰: +134.9° (c 0.545, DMF) Chiral SFC (method SFC-E): R_(t) 4.31min, MH⁺ 529, chiral purity 100%, Enantiomer 3B wherein ¹H NMR (360 MHz,DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.77 (s, 3H) 4.01 (s, 3H) 6.21(d, J=8.1 Hz, 1H) 6.54-6.62 (m, 2H) 6.83 (dd, J=8.6, 2.4 Hz, 1H) 6.91(t, J=1.5 Hz, 1H) 6.94-6.99 (m, 2H) 7.07 (d, J=7.0 Hz, 1H) 7.13 (d,J=1.8 Hz, 1H) 7.35 (d, J=8.1 Hz, 1H) 8.02 (d, J=8.8 Hz, 1H) 8.32 (d,J=2.9 Hz, 1H) 11.87 (br d, J=2.2 Hz, 1H) LC/MS (method LC-A): R_(t) 1.08min, MH⁺ 529 [α]_(D) ²⁰: −167° (c 0.51, DMF) Chiral SFC (method SFC-E):R_(t) 4.63 min, MH⁺ 529, chiral purity 94.7%, Enantiomer 4A wherein ¹HNMR (500 MHz, DMSO-d₆) δ ppm 2.21 (s, 3H) 3.09 (s, 3H) 3.72 (s, 3H) 3.79(s, 3H) 4.01 (s, 3H) 6.20 (d, J=7.6 Hz, 1H) 6.58 (d, J=1.6 Hz, 2H)6.87-6.93 (m, 2H) 6.96 (dd, J=8.2, 1.9 Hz, 1H) 7.02 (d, J=7.6 Hz, 1H)7.12 (d, J=1.9 Hz, 1H) 7.34 (d, J=8.2 Hz, 1H) 7.89 (s, 1H) 8.25 (s, 1H)11.78 (br. s., 1H) LC/MS (method LC-C): R_(t) 3.15 min, MH⁺ 543 [α]_(D)²⁰: +141.8° (c 0.3936, DMF) Chiral SFC (method SFC-C): R_(t) 4.95 min,MH⁺ 543, chiral purity 100%, Enantiomer 4B wherein ¹H NMR (500 MHz,DMSO-d₆) δ ppm 2.21 (s, 3H) 3.09 (s, 3H) 3.72 (s, 3H) 3.79 (s, 3H) 4.01(s, 3H) 6.20 (d, J=7.9 Hz, 1H) 6.58 (s, 2H) 6.88-6.93 (m, 2H) 6.96 (dd,J=8.2, 1.9 Hz, 1H) 7.02 (d, J=7.9 Hz, 1H) 7.12 (d, J=1.9 Hz, 1H) 7.34(d, J=8.2 Hz, 1H) 7.90 (s, 1H) 8.25 (s, 1H) 11.79 (br. s., 1H) LC/MS(method LC-C): R_(t) 3.15 min, MH⁺ 543 [α]_(D) ²⁰: −142.2° (c 0.3909,DMF) Chiral SFC (method SFC-C): R_(t) 6.84 min, MH⁺ 543, chiral purity100%, Enantiomer 5A wherein ¹H NMR (500 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H)3.72 (s, 3H) 3.85 (s, 3H) 4.00 (s, 3H) 6.21 (d, J=7.9 Hz, 1H) 6.58 (d,J=1.3 Hz, 2H) 6.90 (s, 1H) 6.97 (dd, J=8.2, 2.0 Hz, 1H) 7.07 (d, J=7.9Hz, 1H) 7.11-7.17 (m, 2H) 7.34 (d, J=8.2 Hz, 1H) 7.82 (d, J=11.7 Hz, 1H)8.35 (s, 1H) 11.98 (br. s., 1H) LC/MS (method LC-C): R_(t) 3.00 min, MH⁺547 [α]_(D) ²⁰: +136.4° (c 0.28, DMF) Chiral SFC (method SFC-B): R_(t)3.43 min, MH⁺ 547, chiral purity 100%, Enantiomer 5B wherein ¹H NMR (500MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72 (s, 3H) 3.85 (s, 3H) 4.00 (s, 3H)6.21 (d, J=7.9 Hz, 1H) 6.58 (d, J=1.3 Hz, 2H) 6.90 (s, 1H) 6.97 (dd,J=8.2, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.11-7.19 (m, 2H) 7.34 (d,J=8.2 Hz, 1H) 7.82 (d, J=11.7 Hz, 1H) 8.35 (s, 1H) 11.95 (br. s., 1H)LC/MS (method LC-C): R_(t) 3.00 min, MH⁺ 547 [α]_(D) ²⁰: −126.3° (c0.2755, DMF) Chiral SFC (method SFC-B): R_(t) 4.80 min, MH⁺ 547, chiralpurity 98.06%, Enantiomer 6A wherein ¹H NMR (360 MHz, DMSO-d₆) δ ppm2.29 (s, 3H) 3.10 (s, 3H) 3.72 (s, 3H) 3.80 (s, 3H) 4.02 (s, 3H) 6.24(d, J=7.7 Hz, 1H) 6.56-6.59 (m, 1H) 6.59-6.62 (m, 1H) 6.92 (t, J=1.6 Hz,1H) 6.93-6.99 (m, 2H) 7.06 (d, J=7.7 Hz, 1H) 7.13 (d, J=1.8 Hz, 1H) 7.35(d, J=8.4 Hz, 1H) 7.94 (d, J=8.4 Hz, 1H) 8.35 (s, 1H) 11.91 (br s, 1H)LC/MS (method LC-A): R_(t) 1.18 min, MH⁺ 543 [α]_(D) ²⁰: +122.9° (c0.48, DMF) Chiral SFC (method SFC-E): R_(t) 4.15 min MH⁺ 543, chiralpurity 100%, Enantiomer 6B wherein ¹H NMR (360 MHz, DMSO-d₆) δ ppm 2.29(s, 3H) 3.10 (s, 3H) 3.72 (s, 3H) 3.80 (s, 3H) 4.02 (s, 3H) 6.24 (d,J=7.7 Hz, 1H) 6.57-6.59 (m, 1H) 6.59-6.62 (m, 1H) 6.92 (t, J=1.8 Hz, 1H)6.93-7.00 (m, 2H) 7.06 (d, J=7.7 Hz, 1H) 7.13 (d, J=1.8 Hz, 1H) 7.35 (d,J=8.1 Hz, 1H) 7.94 (d, J=8.8 Hz, 1H) 8.35 (d, J=2.2 Hz, 1H) 11.91 (br s,1H) LC/MS (method LC-A): R_(t) 1.22 min, MH⁺ 543 [α]_(D) ²⁰: −120.6° (c0.2755, DMF) Chiral SFC (method SFC-E): R_(t) 4.50 min, MH⁺ 543, chiralpurity 99.35%, Enantiomer 7A wherein ¹H NMR (400 MHz, DMSO-d₆) δ ppm2.30 (d, J=1.5 Hz, 3H) 3.09 (s, 3H) 3.72 (s, 3H) 4.00 (s, 3H) 6.22 (d,J=7.9 Hz, 1H) 6.56-6.60 (m, 2H) 6.91 (t, J=1.7 Hz, 1H) 6.97 (dd, J=8.3,2.1 Hz, 1H) 7.01 (d, J=7.7 Hz, 1H) 7.12 (d, J=2.0 Hz, 1H) 7.22 (d,J=10.1 Hz, 1H) 7.34 (d, J=8.1 Hz, 1H) 8.02 (d, J=7.7 Hz, 1H) 8.37 (s,1H) 11.96 (s, 1H) LC/MS (method LC-A): R_(t) 1.15 min, MH⁺ 531 [α]_(D)²⁰: −163.2° (c 0.435, DMF) Chiral SFC (method SFC-E): R_(t) 4.26 min,MH⁺ 531, chiral purity 100%, Enantiomer 7B wherein ¹H NMR (400 MHz,DMSO-d₆) δ ppm 2.30 (d, J=1.5 Hz, 3H) 3.09 (s, 3H) 3.72 (s, 3H) 4.00 (s,3H) 6.22 (d, J=7.7 Hz, 1H) 6.57-6.61 (m, 2H) 6.92 (t, J=1.8 Hz, 1H) 6.97(dd, J=8.1, 2.0 Hz, 1H) 7.01 (d, J=7.7 Hz, 1H) 7.12 (d, J=2.0 Hz, 1H)7.22 (d, J=10.0 Hz, 1H) 7.35 (d, J=8.4 Hz, 1H) 8.02 (d, J=7.9 Hz, 1H)8.37 (d, J=2.4 Hz, 1H) 11.97 (s, 1H) LC/MS (method LC-A): R_(t) 1.15min, MH⁺ 531 [α]_(D) ²⁰: +166.6° (c 0.5, DMF) Chiral SFC (method SFC-E):R_(t) 3.78 min, MH⁺ 531, chiral purity 100%, Enantiomer 8A wherein ¹HNMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H) 6.29(d, J=7.6 Hz, 1H) 6.60 (br s, 2H) 6.92 (s, 1H) 6.98 (dd, J=8.3, 1.8 Hz,1H) 7.07 (d, J=8.1 Hz, 1H) 7.13 (d, J=1.5 Hz, 1H) 7.36 (d, J=8.1 Hz, 1H)7.54 (d, J=8.1 Hz, 1H) 7.69 (d, J=8.6 Hz, 1H) 8.49 (s, 1H) 8.60 (s, 1H)12.41 (br s, 1H) LC/MS (method LC-C): R_(t) 3.25 min, MH⁺ 567 [α]_(D)²⁰: −119.2° (c 0.2727, DMF) Chiral SFC (method SFC-F): R_(t) 2.64 min,MH⁺ 567, chiral purity 100%, Enantiomer 8B wherein ¹H NMR (400 MHz,DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H) 6.29 (d, J=8.1 Hz,1H) 6.60 (s, 2H) 6.92 (s, 1H) 6.98 (dd, J=8.6, 2.0 Hz, 1H) 7.07 (d,J=8.1 Hz, 1H) 7.13 (d, J=2.0 Hz, 1H) 7.36 (d, J=8.6 Hz, 1H) 7.54 (dd,J=8.6, 1.5 Hz, 1H) 7.69 (d, J=8.6 Hz, 1H) 8.49 (s, 1H) 8.60 (s, 1H)12.40 (br s, 1H) LC/MS (method LC-C): R_(t) 3.25 min, MH⁺ 567 [α]_(D)²⁰: +125.1° (c 0.2455, DMF) Chiral SFC (method SFC-F): R_(t) 3.44 min,MH⁺ 567, chiral purity 100%, Enantiomer 9A wherein ¹H NMR (400 MHz,DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H) 6.26 (d, J=7.9 Hz,1H) 6.55-6.62 (m, 2H) 6.91 (t, J=1.5 Hz, 1H) 6.98 (dd, J=8.4, 2.0 Hz,1H) 7.07 (d, J=7.9 Hz, 1H) 7.13 (d, J=2.0 Hz, 1H) 7.21 (dd, J=8.8, 1.8Hz, 1H) 7.36 (d, J=8.4 Hz, 1H) 7.59 (d, J=8.8 Hz, 1H) 8.07 (d, J=0.9 Hz,1H) 8.55 (s, 1H) 12.29 (br s, 1H) LC/MS (method LC-A): R_(t) 1.20 min,MH⁺ 583 [α]_(D) ²⁰: +130.3° (c 0.555, DMF) Chiral SFC (method SFC-E):R_(t) 3.10 min, MH⁺ 583, chiral purity 100%, Enantiomer 9B wherein ¹HNMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H) 3.99 (s, 3H) 6.26(d, J=7.9 Hz, 1H) 6.56-6.62 (m, 2H) 6.92 (t, J=2.0 Hz, 1H) 6.98 (dd,J=8.1, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13 (d, J=2.0 Hz, 1H) 7.22(dd, J=8.8, 1.8 Hz, 1H) 7.36 (d, J=8.4 Hz, 1H) 7.59 (d, J=8.8 Hz, 1H)8.07 (d, J=0.9 Hz, 1H) 8.55 (s, 1H) 12.30 (br s, 1H) LC/MS (methodLC-A): R_(t) 1.20 min, MH⁺ 583 [α]_(D) ²⁰: −133.2° (c 0.5, DMF) ChiralSFC (method SFC-E): R_(t) 3.50 min, MH⁺ 583, chiral purity 100%,Enantiomer 10A wherein ¹H NMR (360 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72(s, 3H) 3.87 (s, 3H) 3.99 (s, 3H) 6.22 (d, J=7.7 Hz, 1H) 6.55-6.59 (m,2H) 6.88-6.91 (m, 1H) 6.98 (dd, J=8.1, 1.8 Hz, 1H) 7.08 (d, J=7.7 Hz,1H) 7.13 (d, J=2.2 Hz, 1H) 7.21 (s, 1H) 7.34 (d, J=8.1 Hz, 1H) 8.02 (d,J=1.5 Hz, 1H) 8.41 (s, 1H) 12.05 (br s, 1H) LC/MS (method LC-A): R_(t)1.20 min, MH⁺ 613 [α]_(D) ²⁰: +81.4° (c 0.29, DMF) Chiral SFC (methodSFC-E): R_(t) 3.34 min, MH⁺ 613, chiral purity 100%, or apharmaceutically acceptable salt, solvate or polymorph thereof,Enantiomer 10B wherein ¹H NMR (360 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.72(s, 3H) 3.87 (s, 3H) 3.99 (s, 3H) 6.22 (d, J=7.7 Hz, 1H) 6.55-6.60 (m,2H) 6.90 (t, J=1.6 Hz, 1H) 6.98 (dd, J=8.2, 2.0 Hz, 1H) 7.08 (d, J=7.8Hz, 1H) 7.13 (d, J=2.2 Hz, 1H) 7.21 (s, 1H) 7.34 (d, J=8.4 Hz, 1H) 8.01(d, J=1.1 Hz, 1H) 8.41 (s, 1H) 12.08 (br s, 1H) LC/MS (method LC-A):R_(t) 1.20 min, MH⁺ 613 [α]_(D) ²⁰: 99.6° (c 0.261, DMF) Chiral SFC(method SFC-E): R_(t) 3.69 min, MH⁺ 613, chiral purity 100%, Enantiomer11A wherein ¹H NMR (600 MHz, DMSO-d₆) δ ppm 2.50 (s, 3H) 3.09 (s, 3H)3.72 (s, 3H) 4.00 (s, 3H) 6.28 (d, J=7.8 Hz, 1H) 6.56-6.63 (m, 2H) 6.92(br s, 1H) 6.97 (dd, J=8.4, 1.9 Hz, 1H) 7.05 (br s, 1H) 7.07 (d, J=7.9Hz, 1H) 7.13 (d, J=1.9 Hz, 1H) 7.35 (d, J=8.4 Hz, 1H) 7.90 (br s, 1H)8.53 (s, 1H) 12.41 (br s, 1H) LC/MS (method LC-A): R_(t) 1.26 min, MH⁺597 [α]_(D) ²⁰: +81.3° (c 0.3455, DMF) Chiral SFC (method SFC-E): R_(t)2.96 min, MH⁺ 597, chiral purity 100%, and Enantiomer 11B wherein ¹H NMR(600 MHz, DMSO-d₆) δ ppm 2.51 (s, 3H) 3.09 (s, 3H) 3.72 (s, 3H) 4.00 (s,3H) 6.28 (d, J=7.9 Hz, 1H) 6.58-6.60 (m, 2H) 6.92 (t, J=1.8 Hz, 1H) 6.97(dd, J=8.4, 1.9 Hz, 1H) 7.05 (br s, 1H) 7.06 (d, J=7.9 Hz, 1H) 7.13 (d,J=2.1 Hz, 1H) 7.35 (d, J=8.2 Hz, 1H) 7.89 (br s, 1H) 8.53 (s, 1H) 12.37(br s, 1H) LC/MS (method LC-A): R_(t) 1.26 min, MH⁺ 597 [α]_(D) ²⁰:−87.4° (c 0.342, DMF) Chiral SFC (method SFC-E): R_(t) 3.44 min, MH⁺597, chiral purity 100%, or a pharmaceutically acceptable salt, solvateor polymorph thereof.
 8. The method of claim 1, wherein said compoundis:

or a stereoisomer, pharmaceutically acceptable salt, solvate orpolymorph thereof.
 9. The method of claim 1, wherein said compound is:Enantiomer 9A, wherein ¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73(s, 3H) 3.99 (s, 3H) 6.26 (d, J=7.9 Hz, 1H) 6.55-6.62 (m, 2H) 6.91 (t,J=1.5 Hz, 1H) 6.98 (dd, J=8.4, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13(d, J=2.0 Hz, 1H) 7.21 (dd, J=8.8, 1.8 Hz, 1H) 7.36 (d, J=8.4 Hz, 1H)7.59 (d, J=8.8 Hz, 1H) 8.07 (d, J=0.9 Hz, 1H) 8.55 (s, 1H) 12.29 (br s,1H) LC/MS (method LC-A): R_(t) 1.20 min, MH⁺ 583 [α]_(D) ²⁰: +130.3° (c0.555, DMF) Chiral SFC (method SFC-E): R_(t) 3.10 min, MH⁺ 583, chiralpurity 100%, or a pharmaceutically acceptable salt, solvate or polymorphthereof.
 10. The method of claim 1, wherein said compound is: Enantiomer9B, wherein ¹H NMR (400 MHz, DMSO-d₆) δ ppm 3.09 (s, 3H) 3.73 (s, 3H)3.99 (s, 3H) 6.26 (d, J=7.9 Hz, 1H) 6.56-6.62 (m, 2H) 6.92 (t, J=2.0 Hz,1H) 6.98 (dd, J=8.1, 2.0 Hz, 1H) 7.07 (d, J=7.9 Hz, 1H) 7.13 (d, J=2.0Hz, 1H) 7.22 (dd, J=8.8, 1.8 Hz, 1H) 7.36 (d, J=8.4 Hz, 1H) 7.59 (d,J=8.8 Hz, 1H) 8.07 (d, J=0.9 Hz, 1H) 8.55 (s, 1H) 12.30 (br s, 1H) LC/MS(method LC-A): R_(t) 1.20 min, MH⁺ 583 [α]_(D) ²⁰: −133.2° (c 0.5, DMF)Chiral SFC (method SFC-E): R_(t) 3.50 min, MH⁺ 583, chiral purity 100%,or a pharmaceutically acceptable salt, solvate or polymorph thereof. 11.The method of claim 1, wherein said compound is administered in the formof a pharmaceutical composition comprising the compound or thestereoisomer, pharmaceutically acceptable salt, solvate or polymorphthereof, and one or more pharmaceutically acceptable excipients,diluents or carriers.